Chorionic plate vessels as an origin of amniotic fluid neutrophils

The present study was conducted to investigate the potential anatomical source of amniotic fluid neutrophils. Microdissection of neutrophils from the chorioamnion of the fetal membranes and the amnion of the chorionic plates of 10 preterm placentas with acute chorioamnionitis was performed and the genotypes of the neutrophils were compared with those of the mother and fetus using polymerase chain reaction of nine autosomal STR loci. In separate analyses, we reviewed eight cases of fetal autopsies with increased amniotic fluid neutrophils for the presence of neutrophils in the alveoli, and also analyzed the relationship between the amniotic fluid white blood cell (WBC) count and the histological pattern of placental inflammation. The genotypes of all of the neutrophils found in the chorioamnion of the fetal membrane matched those of the mother (n = 10). The genotypes of neutrophils found in the chorionic plate were of mixed maternal and fetal origin (n = 4). In the autopsy series of the fetuses with amniotic fluid WBC (n = 8), only five cases showed neutrophils in the alveolar space, while all the placentas had chorioamnionitis. There was no significant difference in amniotic fluid WBC count between the cases with or without acute membranitis, while among the cases with placental inflammation, those with inflammation of the chorionic plate had a significantly higher amniotic fluid WBC count than both the membranitis-only cases (P < 0.001) and the membranitis and funisitis cases (P < 0.05). These results imply that fetal vasculature at the chorionic plate is the main source of amniotic fluid neutrophils, especially in the cases without funisitis.     

Catecholamines in amniotic fluid as indicators of intrapartum fetal stress

Catecholamines were measured in the amniotic fluid and in the first voided newborn urine obtained from appropriate-for-date infants of term deliveries. Catecholamine values in the amniotic fluid and urine were nearly equal when expressed in terms of creatinine. Significant positive correlations were observed between the amniotic fluid and urine of norepinephrine and epinephrine. In normal cases (n = 32) that underwent uneventful vaginal delivery, the 95% confidence limits for norepinephrine and epinephrine in the amniotic fluid were 1.53 to 2.33 ng/ml and 0.16 to 0.30 ng/ml, respectively. In cases of moderate stress (n = 12), only norepinephrine showed significantly higher values than the normal cases, while in cases of severe stress (n = 12), norepinephrine became more significantly high, and epinephrine was found to be elevated significantly. A significant difference was noted in the incidence of fetal stress between the infants with more than and those with less than 2.30 ng/ml of norepinephrine, the upper limits of the normal 95% confidence limits. However, for epinephrine such a significant difference was not noted. It was concluded that amniotic fluid catecholamines are of fetal origin and reflect fetal sympathoadrenal activity directly, even during labor, and that their level may be a good indicator of fetal condition and stress.     

G, A and M immunoglobulin levels in the parturients' blood serum, umbilical blood of their neonates, and the amniotic fluid in deliveries of prolonged pregnancies

The present experiments were carried out with the aim to determine G, A and M immunoglobulin concentration levels in the blood serum of parturients, umbilical blood of their neonates, and in the amniotic fluid in 17 women with prolonged pregnancies. The diagnosis of prolonged pregnancy was confirmed by the typical after term traits in infants. IgG and IgA were detected in all three environments and the presence of IgM was confirmed in all cases in the blood serum and in 76% of the tested samples of amniotic fluid. IgG concentration in the blood serum of mothers with prolonged pregnancy was lower than in physiological pregnancies. Despite this IgG level in the serum of umbilical blood of the neonates was normal in all cases. The values of IgA and IgM were noted to be normal both in the parturients' blood serum as well as in the umbilical blood. Examination of the amniotic fluid did not reveal significant differences in immunoglobulin levels related to the duration of full term pregnancy compared to prolonged pregnancy.     

Microbiological findings in pregnant women with premature rupture of the membranes

From 30 consecutive cases of premature rupture of the membranes (PROM) and matched controls, specimens from urine, cervix, amniotic fluid, and placenta as well as neonatal nose and throat swabs were investigated bacteriologically and virologically. In addition virus serological investigation was done. Among the PROM cases the anaerobic cervical isolates outnumbered the aerobic ones, and the total number of aerobic as well as anaerobic isolates was less in the control group. The anaerobesB. fragilis andStrept. intermedius were isolated from the cervix, amniotic fluid, or placenta in 23% and 30% of the PROM patients, respectively. None of the controls harbouredB. fragilis, whileStrept. intermedius was isolated from 6.7% of the controls. Group B streptococci were recovered from the mother's cervix in 20% of the PROM patients and in 6.7% among the controls. Four cases of neonatal septicaemia were encountered, and another two cases were clinically suspected, but not microbiologically verified, contributing to a high perinatal mortality rate (17,6%). Of the PROM patients, 27% developed puerperal infection, while none of the control mothers had such complications. The significance of the anaerobic bacteria as well as group B streptococci for the maternal and neonatal outcome in cases of PROM is discussed, and a possible aetiological role of ascending infection in this complication of pregnancy is postulated.     

荧光原位杂交检测在产前诊断中的临床价值

目的探讨运用羊水间期细胞开展荧光原位杂交检测在产前诊断中的临床价值。方法运用FISH技术对110例孕16～24周孕妇的羊水间期细胞进行检测,每例均进行常规染色体核型分析。结果运用FISH法,所有样本均在19h内获得检测结果,除2例羊水培养失败外,所有样本均取得染色体核型分析报告结果。两种方法均检出特氏综合征、21-三体综合征各1例,所有样本的两种方法检测结果均互相符合。常规染色体核型分析有2例异常,因缺乏相应DNA探针。FISH法未能检出。结论运用羊水间期细胞开展荧光原位杂交检测,缩短了诊断时间,缓解了孕妇及家属因等待检测结果所产生的焦虑心情。检测程序简单,结果判定明确,在产前诊断实施过程中具有重要的临床价值。     

Spontaneous bacteriological course of prolonged rupture of the fetal membranes (author's transl)]

Bacteriologic study of amniotic fluid was undertaken in 300 cases of prolonged rupture of the membranes to evaluate the risk of infection without systematic administration of prophylactic antibiotics to the mother. The rate of amniotic contamination was 8.3 per cent within the first 12 hours following rupture of the membranes, it increased with time to reach 52 per cent, after 48 hours. In 90 per cent of the cases, contamination was caused by streptococci and anaerobes, Streptococcus B alone being responsible for 50% of the contaminations. Antibiotics administered to the mother were effective, but they needed a 6 hour-minimum delay to sterilize a contaminated amniotic fluid. Although the overall percentage of contamination was 23 per cent, infection occurred in only 4 per cent of the infants. The outcome was satisfactory in every case. Those results do not seem to discredit the suppression of prophylactic cover antibiotic therapy following the premature rupture of the membranes.     

Concanavalin A reactivity pattern of human amniotic fluid AFP examined by crossed affino-immunoelectrophoresis. A definite test for neural tube defect?

The percentage of alpha-fetoprotein (AFP) not reacting with concanavalin A (con A) was determined by crossed line affino-immunoelectrophoresis in amniotic fluid from 25 pregnancies with neural tube defects (NTD) and other fetal abnormalities giving rise to elevated AFP levels, and from 128 pregnancies with normal outcome. The percentage of con A non-reactive AFP is significantly lower in the presence of fetal abnormalities (mean 3.4 %, range: 0.0–6.3, n = 23, compared to the percentage found in normal pregnancies (mean 17.2 % , range: 6.6–35.8 %, n = 128). In amniotic fluid samples from normal pregnancies, the percentage of con A non-reactive AFP in 84 cases with total AFP levels lying within the 95 % reference interval was not significantly different from the percentage found in 44 cases with levels above the 95 % reference interval. The percentage of non-reactive AFP in fetal serum and cerebrospinal fluid was of the same magnitude as found in amniotic fluids of pregnancies with fetal abnormalities. It is concluded that analysis of the percentage of con A non-reactive AFP by crossed line affino-immunoelectrophoresis is a simple, reliable and apparently diagnostic test for NTD and ether abnormalities with leakage of fetal serum or cerebrospinal fluid into the amniotic fluid. The test should therefore be used in all cases with only marginally elevated AFP levels, so false positives may be avoided.     

Chronic chorioamnionitis displays distinct alterations of the amniotic fluid proteome

Acute chorioamnionitis of infectious origin and chronic chorioamnionitis of immunological origin are two major placental lesions of spontaneous preterm birth with elevated amniotic fluid interleukin-6 and CXCL10 concentrations, respectively. The changes in the amniotic fluid proteome associated with intra-amniotic infection and acute chorioamnionitis are well defined, yet alterations unique to chronic chorioamnionitis remain to be elucidated. This study was conducted to determine those amniotic fluid proteins changing specifically in the presence of chronic chorioamnionitis. Amniotic fluid obtained from acute chorioamnionitis, chronic chorioamnionitis and gestational age-matched controls were analysed by two-dimensional (2D) difference in gel electrophoresis and MALDI-TOF analyses. The type of histological inflammation was used to define each condition in preterm labour cases (n = 125) and term not in labour cases (n = 22), and the amniotic fluid concentrations of interleukin-6, CXCL8, CXCL10 and prostaglandin F(2α) were also measured by specific immunoassays. Among preterm labour cases, 31 differentially expressed proteins were identified in chronic chorioamnionitis cases as compared to both acute chorioamnionitis and control cases. Importantly, glycodelin-A, which maintains maternal tolerance against an allogeneic fetus, was decreased in chronic chorioamnionitis, while haptoglobin was increased. We report the amniotic fluid proteome of chronic chorioamnionitis for the first time, and the findings herein strongly suggest that there is a pathophysiological association between the changes of immunomodulatory proteins in the amniotic fluid and chronic chorioamnionitis, a histological manifestation of maternal anti-fetal allograft rejection.     

产前诊断中羊水嵌合体形成机制的探讨

目的探讨产前诊断中羊水嵌合体形成的机制,便于临床医生在遗传咨询中正确处理羊水培养中出现的嵌合现象。方法 2004年1月至2010年4月对在中山大学附属第一医院胎儿医学中心因各种产前诊断指征就诊的患者行羊膜腔穿刺术取羊水做染色体检查,从中选取羊水染色体为嵌合体的患者穿刺脐血复查染色体,结合两者的结果探讨羊水嵌合体形成机制。结果羊水嵌合体检出33例,其中25例为假嵌合体,8例为真嵌合体,7例真嵌合体羊水结果与脐血结果一致,1例真嵌合体两者结果不一致。结论正确判断羊水培养中出现的真假嵌合体及其风险,对产前诊断和遗传咨询具有重要的意义。     

羊水栓塞救治成功5例的临床研究

目的探讨羊水栓塞（AFE）的有效救活措施。方法回顾性分析我院经治的5例AFE病例，对其临床救治过程进行剖析，总结抢救成功因素。结果剖宫产术中发生的4例急性AFE，采用给氧、大剂量地塞米松、肾上腺素、阿托品、多巴胺等综合急救，全部救治存活。产后以出血为主的1例AFE经纠正DIC、子宫切除后救治存活。结论肾上腺素在救治急性AFE时有使用价值。AFE发生难以纠正的DIC时，切除子宫是重要治疗措施。     

The relationships between thymus, other immune organs and various diseases in children (analysis of 621 cases)

Thymuses, spleens, lymph nodes, tonsils and appendices from 621 autopsy cases were reviewed. The degree of atrophy of these organs might be classified into 3 grades. The main diagnoses of these cases involved more than 130 different diseases, which could be divided into 9 groups: infection, benign tumor, malignant tumor, deformity, amniotic fluid aspiration, fungal infection, autoimmune disease and hemorrhage, etc. Cases of infection with a course less than 5 days showed mild atrophy of the thymus and those cases with a longer course might show moderate or severe degree of atrophy. Cases of benign tumor showed no relationship between the degree of atrophy and the course of the disease. Nevertheless, in cases of malignant neoplasms even there was a history of short course, showed severe atrophy of the thymus. Only mild atrophy was obtained in those patients with deformity, amniotic fluid aspiration and hemorrhage, while severe atrophy was seen in those cases with fungal infection and autoimmune disease. In 81% of the cases, the degree of thymus atrophy was in accordance with those of the other immune organs.     

"M" alpha-1-antitrypsin at term of pregnancy in an African population: concentration and microheterogeneity

For 19 multipars in Ivory Coast alpha-1-antitrypsin (A1AT) has been determined in mother's serum as well as in the cord serum and the amniotic fluid. In all cases the type of A1AT was MM. In 15 cases the pregnancies were normal and full term while four were premature (28th week). The A1AT levels obtained were found to be similar to the literature values and thereby do not seem to be influenced by the ethnic origin. Crossed electroimmunodiffusion analyses demonstrate a distribution of the different fractions of A1AT which is characteristic for each of the above groups of samples analysed. The similarity of the fractions in mother's serum and amniotic fluid indicate a maternal origin for A1AT of the latter. The cord blood shows a very characteristic increase in the peak M6. In the author's experience, they have only noticed such a peak in the blood of patients with primary liver cancer of the phenotype MM.     

Chromosomal mosaicism in amniotic fluid cell cultures

Six cases of chromosomal mosaicism detected in amniotic fluid cultures are described. In five of these there was no evidence of fetal mosaicism. In one case fetal mosaicism was demonstrated but only by the study of fibroblasts since blood cultures showed only normal cells. The implications of amniotic fluid mosaicism are discussed and it is concluded that this usually does not indicate fetal mosaicism. The value of repeated amniocentesis in the diagnosis of fetal mosaicism was demonstrated by findings in three of the cases. It is recommended that amniotic fluid cultures be harvested in situ for chromosome studies and that cytogenetic results be expressed as number of colonies karyotyped rather than as number of cells analyzed.     

荧光原位杂交技术在未培养羊水快速产前诊断中的应用

目的建立运用荧光原位杂交技术（fluorescence in situ hybridization,FISH）检测未培养羊水细胞染色体数目的产前诊断方法。方法对100例妊娠15—26周抽取的羊水未经培养进行FISH检测,均采用13、21、18、X、Y号染色体荧光探针检测,同时行羊水培养染色体核型分析,比较两种检测方法结果的一致性。结果100例产前诊断者中未培养羊水细胞FISH检测均于48h内完成,发现6例21-三体和1例18-三体,与羊水培养染色体核型分析结果一致,后者显示6例21-三体中4例为完全型、2例为易位型21-三体。结论FISH技术与传统的羊水培养染色体核型分析相比较,具有方法快速、简便、准确可靠的特点,但无法完全取代传统的染色体核型分析,应两者结合应用于临床。     

Prenatal diagnosis of genetic disorders.

Three hundred and fifty pregnancies were monitored by transabdominal amniocentesis in the fourteenth to sixteenth week of gestation followed by karyotyping or biochemica assays of cultured amniotic fluid cells and analysis of alpha-fetoprotein in the amniotic fluid supernatant. The pregnancy was interrupted in 36 cases (10%) either becasue of a fetal abnormality or the presence of a male fetus in pregnancies at risk for an X-linked disease. Four chromosomal aberrations were found in 87 pregnancies tested because of advanced maternal age. In 101 pregnancies with a recurrence risk of Down's syndrome, 2 fetuses with an abnormal karyotype were detected. In 11 cases, in which 1 parent was a carrier of a balanced translocation, 2 unbalanced fetal karyotypes were found. Fetal chromosome studies in 43 pregancies at risk for an X-linked disease indicated the presence of a male fetus in 21 cases. Prenatal diagnosis of 11 different metabolic diseases was performed in a total of 34 cases. Microchemical techniques were used to allow completion of the diagnosis of seven different enzyme deficiencies within 9 to 22 days after amniocentesis. Alpha-fetoprotein assay in the amniotic fluid supernatant of 47 pregnancies at risk for an open neural tube defect resulted in the detection of 3 anencephalic fetuses during the second half of pregnancy. The safety and reliability of amniocentesis and the possible effects on the outcome of pregnancy are evaluated. Prenatal diagnosis offers a promising alternative for parents who are at risk of having a child with a genetic disease which can be detected in amniotic fluid or in cultured amniotic fluid cells.     

羊水氨基酸水平与胎儿宫内发育迟缓的关系

目的探讨羊水中氮基酸水平与胎儿宫内发育迟缓(IUGR)的关系.方法采用己硫氢酸苯酯住前衍生法,测定了不同孕周的孕妇60例、IUGR孕妇18例羊水中氨基酸水平.结果随着孕周的增加羊水中总氮基酸水平逐渐下降,至妊娠足月时达最低水平.IUGR组羊水中总氨基酸浓度明显低于正常孕妇组(P<0.05).结论不同孕周羊水中氨基酸含量不同,IUGR时羊水氨基酸水平含量明显降低.     

Rubella in pregnancy: intrauterine transmission and perinatal outcome during a Brazilian epidemic.

Sixty pregnant women with clinical signs of rubella and specific rubella antibodies were studied between January 1999 and December 2002 in order to determine the intrauterine rubella transmission rate and the presence of the virus in amniotic fluid and fetal tissues by nested PCR. Thirty-three patients presented rubella before 12 weeks of gestation and 27 after 12 weeks. Gestational age at the time of acute rubella was determined on the basis of the date of last menstruation and the first trimester ultrasound scan. Thirteen patients with clinical features of rubella before 12 weeks of gestation were submitted to amniocentesis. Three products of conception were analyzed. The presence of the rubella virus was determined by nested PCR. IgM and IgG antibodies were analyzed in neonatal samples at birth and at 3 months of age using a capture immunoassay. Newborn follow-up was based on the presence of congenital rubella syndrome-compatible defects, anti-rubella antibodies, echocardiographic alterations, brainstem evoked response audiometry, and ophthalmological pathology. Five miscarriages and four fetal deaths were observed in the group of patients presenting clinical features before 12 weeks of gestation. IgM antibodies were detected in seven neonates at birth and at 3 months of age. Deafness was observed in three cases and pigmentary retinopathy in one case. Fourteen of the 16 samples (13 amniotic fluid and 3 fetal tissue samples) submitted to virological analysis tested positive. Four fetal deaths, five miscarriages (one with negative virology) and seven newborns with anti-rubella IgM at birth and/or at 3 months age were observed in the group with rubella before 12 weeks of gestation. There were three cases in which virological analysis of the amniotic fluid samples was positive (infected) while the newborn showed no signs of congenital rubella syndrome and anti-rubella IgM were absent. When maternal rubella occurred after 12 weeks of gestation, no fetal or neonatal rubella signs were observed. Eradication of congenital rubella syndrome is possible since vaccination campaigns continue and all services related to the health care of children, adolescents and women have become aware of the significance of the problem and are collaborating. All pregnant women in Brazil should be screened for the rubella antibody and the susceptible group should be vaccinated after giving birth.     

不同孕期羊水间充质干细胞体外培养方法的比较

背景：以往对羊水间充质干细胞的培养多采集孕中期羊水,对妊娠其他阶段羊水进行间充质干细胞培养的研究还很少,特别是系统地对不同孕期羊水和不同培养基进行间充质干细胞培养的效果还未见报道。 目的：比较不同孕期羊水中间充质干细胞体外培养的效果。 方法：采用两种不同细胞培养基分别对60份妊娠时间为15~42周的羊水进行间充质干细胞的分离培养,观察间充质干细胞培养过程中生长状况及间充质干细胞培养成功率;细胞化学染色法检测间充质干细胞的化学性质;MTT法检测传代后间充质干细胞的生长曲线。 结果与结论：采用含体积分数为10%胎牛血清的PRMI-1640培养基和AmnioMAXⅡ complete羊水专用细胞培养基对孕期为37~42周羊水的间充质干细胞培养成功率分别为8%和44%,孕期为21~36周羊水的间充质干细胞成功率分别为20%和60%。而孕期为15~20周的羊水用两种培养基培养成功率为100%。细胞化学染色结果显示：POX、SB(-),ACP、PAS、AENE(+),而NAP有1%为(+),不同孕期染色结果无差异。MTT法测定羊水间充质干细胞的生长曲线呈“S”形。孕期在15~36周的羊水间充质干细胞可以传15代以上仍然旺盛,而孕期为37~42周的羊水间充质干细胞传10代即表现为生长缓慢。结果表明采用AmnioMAXⅡ complete羊水专用细胞培养基可以明显提高晚期羊水间充质干细胞培养的成功率。羊水的最佳采集时间为15~20周。     

An in-vivo study on placental transfer of naproxen in early human pregnancy

BACKGROUND: Naproxen is one of the most common non-steroidal anti-inflammatory drugs used by women of reproductive age. Naproxen is known to be teratogenic in animals. The aim of this study was to investigate the placental transfer of naproxen in the first trimester of human pregnancy, and to determine the amount of the drug in different embryonic compartments. METHODS: Twenty-eight patients who requested surgical termination of pregnancy in the first trimester were given two oral 500 mg doses of naproxen before the surgical procedure. Four biological samples, maternal venous blood, coelomic fluid, amniotic fluid and fetal tissue, were collected from each patient for drug analyses by high performance liquid chromatography. RESULTS: Naproxen was detected in all samples. The mean (+/- SD) concentrations were 69.5 +/- 12.2 microg/ml, 6.4 +/- 2.4 microg/g, 1.85 +/- 1.03 microg/ml and 0.14 +/- 0.11 microg/ml in maternal serum, fetal tissue, coelomic fluid and amniotic fluid respectively. The mean amniotic fluid/maternal drug ratio and fetal/maternal drug ratio were 0.002 (range 0.0005-0.0064) and 0.092 (range 0.022-0.155) respectively. There was a positive correlation between the fetal drug concentration (r = 0.59, P = 0.001), amniotic fluid drug concentration (r = 0.47, P = 0.013), amniotic fluid/maternal ratio (r = 0.536, P = 0.003) and fetal/maternal ratio (r = 0.72, P < 0.001) with advancing gestational age. CONCLUSIONS: Although naproxen can cross the placenta readily in the first trimester of human pregnancy, only a small amount was present in fetal tissues. Since there is no information on whether this small amount of naproxen would be teratogenic or not, women of reproductive age who are taking naproxen regularly should be warned of the possible fetal side-effects.     

Capnocytophaga species: a cause of amniotic fluid infection and preterm labour

Subclinical amniotic fluid infection and subsequent preterm labour may occur with intact membranes. We report two cases of subclinical amniotic fluid infection with intact membranes presenting in preterm labour. Capnocytophaga species, fastidious Gram-negative bacilli normally found in oral flora, were isolated in pure culture from amniotic fluid obtained by transabdominal amniocentesis. The distinctive microbiological features and spectrum of infections associated with Capnocytophaga species, and the importance of recognition of subclinical amniotic fluid infection as a cause of preterm labour, are discussed.