Differential diagnosis of Thrombocytopenia newly developed during pregnancy / 대한산부인과학회지

OBJECTIVE: Gestational thrombocytopenia has a mild course as a common problem during pregnancy, whereas idiopathic thrombocytopenic purpura (ITP) presents with a chronic or moderate to severe course. Very few studies have been conducted so far to discriminate between the two diseases. This study was aimed to identify factors predictive of the subsequent development of ITP among pregnant women presenting with thrombocytopenia. METHODS: From January 1999 to June 2005, a total of 58 pregnant women newly diagnosed with thrombocytopenia were recruited for the study. Among them, 33 were finally diagnosed with gestational thrombocytopenia and the other 25 with ITP. The clinical factors for each of ITP and neonatal thrombocytopenia were evaluated, and final outcomes were also described. RESULTS: On multivariate analysis, thrombocytopenia diagnosed before 28 completed weeks and platelet count of less then 50 x 10(9)/L at the time of its presentation were found to be independently predictive of ITP (p<0.001 and p=0.004 respectively). In addition, platelet count of less then 20 x 10(9)/L at nadir during pregnancy was a significant risk factor for neonatal thrombocytopenia (p=0.013). CONCLUSION: The onset time of thrombocytopenia and platelet count at its presentation remain useful parameters to discriminate ITP from gestational thrombocytopenia. These findings may help in allowing an appropriate antenatal care and postpartum follow up.

Clinical Outcome of Pregnancy-associated Aplastic Anemia Treated with Supportive Anagement / 대한주산의학회잡지

OBJECTIVE: Pregnancy-associated aplastic anemia remains a rare occurrence. The aim of this study was to examine the maternal and fetal outcomes of pregnancy-associated aplastic anemia treated with supportive care. METHODS: From January 1995 to December 2004, a total of 14 women newly diagnosed with pregnancy-associated aplastic anemia were recruited for the study. RESULTS: Eleven (78%) of the 14 women were diagnosed with pregnancy-associated aplastic anemia during the second or third trimester. There were eight severe cases; three of which were diagnosed at the initial presentation. All 14 women had conservative management with transfusions but not specific immunological or hormonal therapies during pregnancy. Blood transfusions were performed prenatally in seven mothers and perinatally in 13. Of the 12 patients eligible for follow-up, one achieved complete remission and another eight showed partial remission after delivery. During the follow up period, there was no case of maternal-fetal death in our series. The pregnancies were continued uneventfully in most cases. CONCLUSIONS: This study demonstrated favorable maternal and neonatal outcomes with transfusion support alone for pregnancy-associated aplastic anemia. Therefore, pregnancy continuation with meticulous blood support should be considered, rather than therapeutic termination, for women with pregnancy-associated aplastic anemia.

The Outcome of Pregnancy Combined with Idiopathic Thrombocytopenia Purpura and the Effect of Pregnancy on the Severity of This Disease / 대한주산의학회잡지

OBJECTIVE: The incidence of idiopathic thrombocytopenic purpura (ITP) is greatest in female during their childbearing years, so the concurrence of pregnancy and ITP is not unusual. Numerous studies have examined the outcomes of newborns, whereas fewer studies have been conducted with regard to the morbidity of obstetric patients with ITP. This study was aimed to find the outcome of pregnancy combined with ITP and the influence of the pregnancy on the severity of this disease. METHODS: From January 1996 to December 2005, a total of 62 pregnant women with ITP and their 73 deliveries were recruited for the study. Among them, 38 were diagnosed with ITP during pregnancy and the other 24 had pre-existing ITP before pregnancy. RESULTS: The severity of thrombocytopenia was exacerbated during pregnancy, but recovered to a level of non-pregnant period after delivery in most cases. The outcome of pregnancy of all the patients was uneventful except each one case of fetal demise at 35 gestational weeks and preterm delivery at 30 gestational weeks. One patient suffered from multiple subdural hemorrhage during pregnancy, which was spontaneouly recovered. Twenty newborns (27.8%) had transient congenital thrombocytopenia and 18 of them required treatment for hemostatic impairment. CONCLUSION: For women with ITP, Pregnancy can affect the severity of ITP, but life-threatening complication was almost lacking. Although, in not a few cases, there may need to treat both mothers and infants to raise their platelet counts, most mothers with ITP can proceed with their pregnancies and delivery healthy infant without complication.

Prenatal Diagnosis of Congenital Diaphragmatic Hernia in a Fetus with 46,XY/46,X,-Y,+der(Y)t(Y;1)(q12;q12) Mosaicism: A Case Report

Congenital diaphragmatic hernia (CDH) is often associated with major anomalies and chromosomal abnormalities. Chromosomal abnormalities are usually detected in 9.5% to 34% of fetuses with CDH prenatally diagnosed and the defect has also been reported in association with multiple syndromes such as Pallister-Killian syndrome, Fryns syndrome, Di George syndrome and Apert syndrome. Among the chromosomal abnormalities associated with CDH, trisomy 21, 18, and 13 are most common. Association with complex chromosomal aberrations such as mosaicism has also been reported. However, CDH presented in a fetus with Y-autosome translocation is extremely rare. Herein, we reported a case of fetus with 46,XY/46,X,-Y, +der(Y)t(Y;1)(q12;q12) mosaicism who presented with CDH diagnosed by ultrasonography at 19 weeks' gestation.

Cytogenetic Analysis in Spontaneous Abortion / 대한주산의학회잡지

OBJECT: To analyze the frequency and the types of chromosomal abnormalities in conceptus of spontaneous abortion method: 81 cases of products of conception aborted spontaneously were collected From Nov. 1999 to May, 2004 in the cytogenetic laboratory at Kangnam St. Mary's Hospital, Catholic University Medical College. The cytogenetic results,along with clinical information including gestational age at the time of the miscarriage and maternal age, were compiled in a database. RESULTS: Cytogenetic results were obtained from 76 cases (93.8%), of which 33 cases (43.4%) showed chromosome abnormalities. Of 33 chromosomal abnormalities, 12 cases (36.4%) showed monosomy X, including one case of 46,X,+7 and the other case of 44,X, t(13q14q). Autosomal trisomy was also detected in 12 cases (36.4%). The most frequent autosomal trisomy was that of chromosome 18 (4 cases), which occurred exclusively in old maternal age (37.8+/-7.5 years old). Mosaicism was found in 2 cases. Triploidy was identified in 4 cases. Structural abnormalities were identified in 3 cases. CONCLUSION: This study showed that monosomy X and trisomy were the most common chromosomal abnormalities in spontaneous abortion, and autosomal trisomy was dependant on maternal age.

Secondary acute myeloid leukemia after platinum based chemotherapy for ovarian cancer / 대한산부인과학회지

Among chemotherapeutic regimens used for advanced ovarian cancer, platinum-based combination chemotherapy remains a mainstay of the treatment of advanced ovarian cancer, providing significant response rates and survival benefits. However, with widespread use of long-term chemotherapy in treating ovarian cancer, emergence of secondary leukemia has become medical concern as one of the most unfavorable late complications. Depending upon the type, duration, and dosage of previous chemotherapy, the risk of developing acute myeloid leukemia has been estimated to be between 2% and 10%. Moreover, the frequency of this complication might increase as the survival in patients with ovarian cancer undergoing chemotherapy continues to increase with developing therapeutic options. Recently, we experienced a case of secondary acute myeloid leukemia developing 3.5 years after platinum-based chemotherapy. In this report, clinical course of the patient and contributing factors for the secondary leukemia were presented.

A case of Vascular Anastomoses in Dichorionic Diamniotic-Fused Placentas Resulting in "Pseudo" Twin-to-Twin Transfusion Syndrome / 대한산부인과학회지

Virtually all monochorionic twin placentas contain vascular connections between the circulatory domains of each twin. In contrast, vascular anastomoses are generally thought not to occur in fused, dichorionic placentation. In the twin-to-twin transfusion syndrome (TTTS) one twin is preferentially perfused by blood from the cotwin via unbalanced placental vascular anastomoses. This vascular shunting results in twins born with discrepant weights, colors and hemoglobins. When one of the TTTS criteria was not present, the pregnancy was defined as "pseudo" TTTS. In pregnancies complicated by "pseudo" TTTS indicate that small twins have abnormal cord insertion more frequently than large twins. We present a case of vascular anastomoses in dichorionic diamniotic-fused placentas resulting in "pseudo" TTTS with a brief review of the literatures concerned.

The length of active labor in women with vaginal birth after cesarean section compared with nulliparas and multiparas / 대한산부인과학회지

OBJECTIVE: Despite the general information of vaginal birth after cesarean section (VBAC), little is known about the duration of active labor in women attempting VBAC. The aim of this study was to compare the time length of active labor in women attempting VBAC compared with nulliparas or multiparas, and then, provide further insight for better management of labor. METHODS: From January 1999 to December 2003, a total of 444 patients with VBAC were entered into the study. Women with two or more history of caesarean section or previous vaginal delivery were all excluded from the study. Time length of active labor in these patients was compared with 335 nulliparas and 218 multiparas consecutively visiting our unit for delivery in 2003. RESULTS: For patients with VBAC, the duration of active and second phase were 184.8+/-115.7, and 25.1+/-15.2 minutes, which was significantly shorter than nulliparas (p<0.05), but longer than multiparas (p<0.05). To eliminate confounding factors affecting the duration, 374 women without use of vacuum, oxytocin, or epidural anesthesia were selected for analysis. Consistently, the time length of active and second phase in women with VBAC was significantly shorter than in nulliparas, but longer than in multiparas (p<0.001) except for active phase compared with nulliparas (p=0.295). CONCLUSION: With regard to the time length of active labor, women attempting VBAC were in the intermediate group when compared with nulliparas or multiparas. Understanding of this unique time interval in women attempting VBAC may help in allowing an appropriate management for labor process.

Diagnosis of Cervical Neoplasia Using Immunohistochemical Staining of p16(INK4A) / 부인종양

OBJECTIVE: There is need for more objective diagnostic parameters to identify cervical dysplastic or neoplastic cells. So, we examined the p16(INK4A) expression in the cervical tissues to evaluate the value of p16(INK4A) as a diagnostic parameter. METHODS: We examined the p16(INK4A) expression by immunohistochemical staining in normal cervical tissues (n=3), preneoplastic lesions (n=6), carcinoma in situ (CIS, n=5), and invasive carcinomas (n=5) of the cervix, which were selected randomly by H and E staining from the archives of formalin-fixed and paraffin-embedded tissues and we also examined the status of human papillomavirus (HPV) infection in the same tissues. RESULTS: The positive rates of p16(INK4A) expression was significantly higher in all abnormal cervical tissues including subclinical papillomavirus infection (SPI), dysplasia, CIS, and invasive carcinoma than in normal cervical epithelium (p=0.001). Despite the strong expression of p16(INK4A) in the area of CIS, no expression of p16(INK4A) was observed in the area of normal epithelium in the vicinity of CIS. 11 cases among 19 cases of examined tissue samples were tested for HPV infection. Seven of them showed positivity for HPV DNA. CONCLUSION: We herein demonstrated that p16(INK4A) would be a sensitive and specific marker for the abnormal cervical cells in tissue sections. This approach will help to reduce interobserver variations in the histopathologic interpretation of cervical biopsy specimens.

Clinical significance of Urinary protein fractional analysis in Severe preeclampsia / 대한산부인과학회지

OBJECTIVE: To determine whether severity of proteinuria or urinary protein fractional analysis correlates with adverse maternal and fetal outcomes in women with severe preeclampsia. METHODS: Thirty-six women diagnosed of severe preeclampsia from January, 2002 to April, 2003 were studied. The correlation between proteinuria or urinary albumin fraction, and maternal mean arterial pressure, neonatal birth weight, 1 minute apgar score were analyzed statistically. Thirty-six patients were divided into two groups according to the pattern of urinary protein fraction. One group was a selective proteinuria group if the albumin fraction was over 70%, and another was a non-selective proteinuria group if the fraction was below 70%. The maternal and neonatal outcomes were compared between the two groups. RESULTS: Significant positive correlation was observed between proteinuria and mean arterial pressure, between urinary albumin fraction and neonatal birth weight. Negative correlation was significantly present between proteinuria and neonatal birth weight, 1 minunte apgar score, between proteinuria and albunin fraction. Increased proteinuria, higher mean arterial pressure, higher serum uric acid level, lower creatinine clearance, lower neonatal birth weight, and lower 1 minunte Apgar score were observed in the non-selective proteinuria group than those in the selective proteinuria, although there was no statistical significance. CONCLUSION: With increasing proteinuria and decreasing albumin fraction, there is increased risk of adverse maternal and fetal outcome. Proteinuria fractional analysis by electrophresis might provide useful information regarding the prediction of pregnancy outcomes.

The Impact of Previous Cervical Dilatation on the Duration of Vaginal Birth after Cesarean Delivery / 대한주산의학회잡지

OBJECTIVE: This study examined the effect of a maximal cervical dilatation prior to their primary cesarean birth on the time length of labor in women attempting vaginal birth after cesarean section (VBAC). METHODS: From January 2000 to Jun 2005, a total of 526 patients with VBAC were entered into the study. Patients were classified into 4 groups according to the maximal cervical dilatation on the prior cesarean birth: Group 1 (0~3 cm), Group 2 (4~7 cm), Group 3 (8~9 cm), and Group 4 (10 cm). RESULTS: An analysis of the duration of labor showed that the time length of active phase was significantly shorter in each of Group 3 (147.3+/-103.9 minutes) and 4 (155.2+/-104.2 minutes) than in Group 1 (192.9+/-126.0 minutes) or 2 (195.1+/-148.2 minutes) (p or =8 cm than those without. Study results indicate that prior cervical dilatation may affect the labor time of subsequent VBAC. The proper understanding of the association between prior cervical dilatation and labor time on a subsequent VBAC may be useful for better management or further intervention in the setting of VABC.

Is Selective Iron Supplementation Needed During Gestation? / 대한산부인과학회잡지

OBJECTIVE: It is controversial whether routine or selective iron supplementation during gestation is needed. The aim of this study is to evaluate whether screening with serum ferritin during the first trimester of pregnancy could be identify women who need prophylactic iron supplementation. METHODS: According to the serum ferritin level of cut-off point of 30 microgram/L during the first trimester of pregnancy and the presence of iron supplementation during gestation, the subject was divided into 4 group that were as follows: serum ferritin level of 30 microgram/L and iron- supplemented group (n=40), serum ferritin level of >30 microgram/L and non iron-supplemented group (n=15). Maternal serum ferritin, TIBC, iron, hemoglobin, hematocrit, red cell indices (MCV, MCH, MCHC, RDW) were measured before 14 weeks of gestation and after 34 weeks of gestation. RESULTS: All hematologic and biochemical markers indicated more severe anemic status in the late pregnancy than during the first trimester of pregnancy. The effect of iron supplementation was profounder on the pregnant woman whose ferritin levels were below 30 microgram/L during the first trimester of pregnancy. Regardless of iron supplementation, the group (ferritin >30 microgram/L during the first trimester of pregnancy) showed relatively higher ferritin level in late pregnancy. CONCLUSION: The screening with serum ferritin level of cut-off point of 30 microgram/L during the first trimester of pregnancy may be useful to identify women who need prophylactic iron supplementation.

Patterns of Hyrtl's Anastomosis and their Effects on Obstetric Outcomes / 대한산부인과학회잡지

OBJECTIVE: This study was conducted to investigate the presence and pattern of Hyrtl anastomosis, and to examine the effect that each type has on obstetrical outcome. METHODS: This study was carried out from January of 2001 to May of 2003 on 904 randomly selected patients who had given birth by Cesarean section or vaginal delivery at Catholic University Holy Family Hospital. Immediately proceeding delivery, barium sulfate was infused into the umbilical artery. This was followed by umbilical artery dissection to grossly confirm Hyrtl anastomosis. In order to evaluate the effect that each type of anastomosis has on obstetrical outcome, we retrospectively examined chart for birth weight, Apgar score, results of cord blood gas analysis, obstetrical complications, etc. RESULTS: Out of the 904 cases studied, 99.7% possessed Hyrt's anastomosis with frequency according to type as follows: 50.8% oblique, 48.0% transverse, 1.8% of anastomosis between one umbilical artery and the branch of the other, 1.6% of fusion of branches of each umbilical artery, 0.6% of anastomosis represented by two separate vessels between umbilical arteries, 0.1% of anastomosis between branches of each umbilical artery, 0.1% lacking anastomosis, and 0.2% of a single umbilical artery. In terms of obstetrical outcome, there was no significant difference for term and preterm infants in the frequency of type of Hyrtl's anastomosis according to birth weight, maternal age, Apgar score, obstetrical complications, etc. CONCLUSION: In an attempt to verify the presence and type of Hyrtl anastomosis, 8 types of Hyrtl's anastomosis were found, of which the oblique and transverse variety were the most common. There was no significant difference in the types of Hyrtl's anastomosis in relation to obstetrical outcome. Frequency was also similar with oblique and transverse types being the most common. Further studies detailing anatomical features of and blood flow/volume through the anastomosis are required in order to better understand the effects of Hyrtl's anastomosis on obstetrical outcome.

Labor Induction at Term with Different Regimens of Misoprostol / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to compare the efficacy and the safety of two different dose regimens of intravaginal misoprostol for labor induction. METHODS: Sixty-seven consenting women with maternal or fetal indication for labor and no prior cesarean delivery were randomly assigned to receive either vaginal misoprostol 100 microgram or 50 microgram every 4 hours with maximum 6 doses. Uterine contraction, abnormal fetal heart rate pattern, mean time from induction to vaginal delivery, apgar score, and meconium staining were compaired. RESULTS: In two groups, dermographic characteristics and the average number of doses of misoprostol used were simliar. The group who were administered 100 ug misprostol showed shorter average time interval from induction to delivery. Both groups showed similiar result in terms of tachysystole, abnormal fetal heart rate tracing, meconium passage, fetal weight, low 1-min and 5-min Apgar scores, and admission to the neonatal intensive care unit. CONCLUSION: The optimal misoprostol dose to induce labor in term pregnant women has not been established. We also could not find any difference between two groups (100 microgram or 50 microgram every 4 hours) in time interval from induction to delivery and neonatal outcomes.

Midtrimester Amniotic Fluid Levels and Each Ratio of Activn A, Inhibin A and B in Down's Syndrome and Other Complicated Pregnancies / 대한산부인과학회잡지

OBJECTIVE: To evaluate the amniotic fluid levels and each ratio of activin A, inhin A and B in Down's syndrome and other complicated pregnancies. METHODS: This study was performed in 71 women who had undergone a midtrimester amniocentesis with the clinical indications and whose pregnancy outcome was retrospectively determined. Ten Down's syndrome, 15 complicated pregnancies including, preeclamsia, gestational diabetesis mellitus, preterm labor, intrauterine growth restriction (IUGR), and 46 noncomplicated pregnancies with normal chromosome were included in this study. Amniotic fluid activin A, inhibin A and B were measured using enzyme linked immunosorbent assays (ELISA). Statistical analysis was performed with Mann-Whitney U test and regression analysis. RESULTS: There were significant positive correlation (r=0.277, p=0.011) between the ratio activin A/ inhibin B level and maternal age and significant positive correlation (r=0.261, p=0.015) between maternal age and the ratio inhibin A/inhibin B level. There were also significant positive correlation (r=0.202, p=0.045) between gestational weeks and inhibin A levels and significant positive correlation (r=0.474, p<0.001) between gestational weeks and inhibin B levels. Amniotic fluid inhibin A and inhibin B levels were significantly (p<0.05) decreased in Down's syndrome compared with the normal chromosomal groups but there was no difference in the ratio activin A/inhibin A and in the ratio activin A/inhibin B between these groups. The amniotic fluid levels of activin A, inhibins (A and B), each ratio of complicated pregnancies groups with normal chromosome was not significantly different from those of uncomplicated pregnancies with normal chromosome. CONCLUSION: This study revealed that amniotic inhibin levels were significantly decreased in Down's syndrome. But, activin A and each ratio were unchanged. The results suggest that activin A may be relatively decreased in each case of Down's syndrome.

A Case of Vesicoallantoic Cyst Diagnosed by Prenatal Ultrasonogram Imaging / 대한주산의학회잡지

Fetal vesicoallantoic cyst is extremely rare anomaly of umbilical cord. This disorder is thought to be a remnant of the extraembryonic portion of the allantois and incomplete obliteration of the vesico-allantoic lumen results in a wide spectrum of urachal anomalies. A 23-years-old primigravida was referred to our hospital at 18 weeks of gestation age because of fetal abdominal cystic mass in basal part of umbilicus. On the ultrasonographic examination, the cystic mass appeared to be in direct communication with the urinary bladder markedly distended. The cytogenetic study by amniocentesis showed normal 46, XY. The fetus was dead at 21 weeks of gestation age. The results of autopsy showed multiple anomalies. We report this rare case with review on similar anomalies of the allantois.

Clinical Study for Intrauterine Fetal Death / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to evaluate intrauterine fetal death and to elucidate the etiology of intrauterine fetal death. METHODS: This is a clinical study of 153 cases of fetal death in utero (FDIU) among 11,866 deliveries at Holy Family Hospital during Mar. 1995 to Feb. 2002. RESULTS: 1. The average incidence of FDIU was 1.29%. 2. The age distribution of mother with FDIU was between 16 to 45 year old and was highest in the 25 to 29 year old age group (45%). 3. The parity of mothers with FDIU was the highest in nulliparous group (58.5%) and there was a decreased tendency with high parity. 4. There were 80 cases (52.1%) with previous history of abortion and 8 cases (5.2%) with previous history of FDIU. 5. The highest incidence rate of FDIU was shown at 20~24 weeks of gestation (36.6%) and in the fetus weighted less than 1,000 gm (56%), and the sex ratio of male versus female fetus was 1.29:1. 6. The mode of delivery FDIU was labor induction (54.3%), laparotomy (15.2%), spontaneous labor (30.4%). The indication for laparotomy were placental abruption, previous cesarean section state, twin and maternal death. As the gestational age and fetal weight decreased, the mean time interval from start of labor induction to fetal expulsion and the variety of labor induction became increase. 7. The etiology factors of FDIU was unexplained causes (44%), congenital anomaly (17%), cord complication (14%) in order. CONCLUSION: The proper antenatal care should be taken of fetuses on the basis of risk factors of antepartum and intrapartum so that unnecessary intrauterine fetal death might be able to be prevented.

The Expression of Vascular Endothelial Growth Factor, Platelet-Derived Growth Factor and Intercellular Adhesion Molecule in Severe Preeclamptic Placenta / 대한산부인과학회잡지

OBJECTIVE: To determine the expression of vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and intercellular adhesion molecule (ICAM-1) in placenta from pregnancies complicated by severe preeclampsia and normal pregnancies. METHODS: Placental tissues were obtained from 10 normotensive pregnancies (control group) and 20 severe preeclamptic pregnancies (preeclamptic group). Immunohistochemical staining of placental tissue was used to determine tissue expression of VEGF, PDGF and ICAM-1. The intensity of staining was evaluated by scoring as 0, 1, 2 and 3. RESULTS: Immunolocalization of VEGF and PDGF was significantly observed in the syncytotrophoblast with less intense staining in intravillous stromal cells and intravillous endothelial cells of fetal vessels in preeclamptic group. There were no differences in immunolocalization of staining in control group. Intensity of VEGF and PDGF immunostaining in syncytotrophoblast was significantly increased in preeclamptic group. However, immunolocalization and the intensity of ICAM-1 staining were not significantly different in both groups. CONCLUSION: The expression of VEGF and PDGF in the syncytotrophoblast was significantly up-regulated in severe preeclamptic placenta. These up-regulation of VEGF and PDGF might reflect that placental ischemia and hypoxic state in severe preeclampsia induce VEGF and PDGF in the syncytotrophoblasts of placenta. However the unchanged pattern of ICAM-1 expression in severe preeclampsia suggests that ICAM-1 is unlikely to be a factor by which the adverse pregnancy outcome arises in severe preeclampsia.

Detection of p16(INK4A) in the Mixed Cell Populations of Normal Peripheral Blood Mononuclear Cells and Cervical Cancer Cell Lines / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Human papilloma viruses (HPVs) play a central role in the pathogenesis of neoplastic lesions of the uterine cervix. The viral oncoprotein HPV E6 degrades the p53 protein, and the HPV E7 protein inactivates pRB and increases the expression of the CDK inhibitor, p16(INK4A). We investigated the usefulness of p16(INK4A) as a biologic marker for the cervical dysplastic and neoplastic cells. MATERIALS AND METHODS: We examined the expression of p16(INK4A) and cytokeratin in a mixed population of normal peripheral blood mononuclear cells (PBMC) and the cervical cancer cell lines (HeLa, SiHa, and CasKi) using flow cytometry. RESULTS: The DNA indices of the HeLa, SiHa and CasKi cell lines were 1.89, 1.53 and 1.75, respectively, indicating that these cells are aneuploid cells. Furthermore, the positive rate of p16(INK4A) expression was 86.7% for the HeLa mixed population, 85.6% for the SiHa mixed population, and 92.2% for the CasKi mixed population. According to the FL3A vs FL3W histogram, electrical gating of the HeLa, SiHa and CasKi mixed populations showed the expression levels of both cytokeratin and p16(INK4A) to be identical, at 86.6%, 84.8% and 85.0%, respectively. These findings revealed that almost all cells selected through electrical gating were cervical cancer cells originating from the epithelium and which expressed cytokeratin and p16(INK4A). On the other hand, when each mixed population was electrically gated for normal PBMC, we found that the PBMCs expressed neither cytokeratin nor p16(INK4A). CONCLUSION: Using flow cytometry, we observed the enhanced expression of p16(INK4A) in cervical cancer cell lines. These RESULTS suggest the usefulness of p16(INK4A) for the selective detection of cervical dysplastic and cancer cells in the liquid-based samples, which are taken from the cervices and contaminated with blood and stromal cells.

Detection of Apoptosis by the Stainings of Annexin V, Propidium Iodide and Cytokeratin in OVCAR-3 Ovarian Cancer Cell Line / 대한산부인과학회잡지

OBJECTIVE: This study was designed to estimate the chemosensitivity by a quantitative evaluation of the apoptotic cell fractions using flow cytometry. METHODS: The OVCAR-3 cells were exposed to 20 nM or 30 nM taxol for 0 (control), 24 and 48 hours, then removed the taxol contained media, and cultured further with fresh media without taxol. (1) Fluorescein isothiocyanate-conjugated Annexin V (Annexin V-FITC) and propidium iodide (PI) were added to one test tube to detect the apoptotic cell fractions and at the same time, PI was added to the other tube to stain the DNA. (2) Annexin V-FITC and cytokeratin (clone CAM5.2 and MNF116) were added to the test tube. They were fixed and permeabilized with 1% paraformaldehyde solution and 100% methanol. They were then incubated with phycoerythrin (PE)-conjugated goat anti-mouse immunoglobulin G (GAM IgG1-PE or GAM IgG2a-PE) and sequentially stained with PI for DNA. All the stained cells were analyzed by a FACScan flow cytometer. RESULTS: (1) After treatment of 20 nM or 30 nM of taxol, G2M arrest was observed in both of treatment groups, which increased with time. (2) The G0G1 sub-fraction indicative of apoptosis increased with increase of culturing time from 24 hrs to 48 hrs. (3) The early apoptotic cell fraction with positive annexin V-FITC and negative PI increased with increase of culturing time. (4) In cells stained sequentailly with annexin V-FITC, cytokeratin (CAM5.2 and MNF116), and PI after 30 nM taxol treatment, the early apoptotic cell fractions increased with increase of culturing time. However, their extent was somewhat lower than those observed by positive annexin V-FITC and negative PI in cells treated with 20 nM of taxol. CONCLUSION: The results of sequential stainings with annexin V-FITC, cytokeratin, and PI were consistent with the those of annexin V-FITC and PI with parallel DNA staining. Our results suggested that the level of apoptosis detected by flow cytometry could be a marker of chemosensitivity which could select the sensitive anti-cancer agents before administration to gynecologic cancer patients.