Damir Roje

Maternal and Neonatal Effects of Substance Abuse during Pregnancy: Our Ten-year Experience

Purpose The aim of the study was to assess perinatal outcome of pregnancy burdened with maternal addiction in comparison with an unselected population from a European transition country. Materials and Methods Data on pregnancies complicated by illicit drug abuse (n = 85) managed during a 10-year period (1997 - 2007) at Split University Hospital were analyzed. Data on the type of drug, course of gestation and labor, and on perinatal outcome were considered. Data on all non-dependence pregnancies recorded during the study period were used as a control group. Results During the study period, there were 85 dependence-complicated pregnancies (0.2%). Use of heroin alone during pregnancy was recorded in 51 women (50%), methadone alone in 6 (7%), and a combination of heroin and methadone in 9 (11%). Premature delivery was significantly more common in the group of pregnant addicts (21% vs. 6%); 49% of pregnant addicts were carriers of hepatitis C virus (HCV) and 14% of hepatitis B virus (HBV). Neonatal abstinence syndrome developed in 61 infants (7%) born to addicted mothers. There were 4 cases (4.6%) of early neonatal death; 7 neonates had 5-minute Apgar score ≤ 7 (8%); 29 neonates had low birth weight for age (33%); and 7 neonates had congenital anomalies (8%). The risk of various congenital anomalies was 3-fold in the group of children born to addicted mothers. Conclusion Addiction pregnancies present a small but high-risk group according to perinatal outcome. Appropriate obstetric and neonatal care can reduce the rate of complications in these pregnancies and improve perinatal outcome.

Trophoblast apoptosis in placentas from pregnancies complicated by preeclampsia.

Aims: To assess trophoblast apoptosis separately in the cytotrophoblast, syncytiotrophoblast, total villous trophoblast, syncytial knots and syncytial knot formation, and to investigate the expression of apoptotic factors Fas ligand (FasL), Bcl-2 and proliferation marker Ki-67 in the trophoblast of placentas from preeclamptic patients. Methods: The study included placental samples from 25 preeclamptic and 25 normal pregnancies. For the detection of apoptosis and proliferation, antibody M30 and antibody against Ki-67 antigen were used. Expression of FasL and Bcl-2 was assessed using semi-quantitative HSCORE method. Syncytial knots were expressed as the number of syncytial knots per individual villus and as the total number of syncytial knots in each placental sample. Results: Trophoblast apoptosis, number of syncytial knots per individual villus and the total number of syncytial knots in each placental sample were significantly higher in preeclamptic placentas than in control group placentas. FasL expression was significantly less, and Bcl-2 expression significantly greater in the villus trophoblast among the study subjects compared with controls. There was no difference in the trophoblast proliferation between groups. Conclusion: Our findings might suggest that increased apoptosis and syncytial knot formation combined with reduced FasL expression could be involved in pathophysiological mechanisms of preeclampsia.

Apoptosis, proliferation and Fas ligand expression in placental trophoblast from pregnancies complicated by HELLP syndrome or pre-eclampsia.

Objective. To investigate apoptosis, proliferation and Fas ligand expression of placental trophoblast in the hemolysis, elevated liver enzymes, low platelets (HELLP) syndrome and in pre‐eclampsia (PE), and to compare this with normal pregnancies. Design. Prospective study. Setting. University hospital in Croatia. Sample. Placentae from women with HELLP syndrome (n=10), PE (n=10) and normal pregnancies (n=10). Methods. The HELLP syndrome was diagnosed with platelets <100×109/L, aspartate aminotransferase (AST) and alanine transaminase (ALT) >70U/L and lactic acid dehydrogenase (LDH) > 600U/L. Pre‐eclampsia was diagnosed at blood pressure >140/90mmHg, with proteinuria >300mg/L/24hours. For detection of apoptosis and proliferation in villous trophoblast, antibodies M30 and Ki‐67 were used. Expression of Fas ligand was assessed using immunohistochemistry and the semiquantitative HSCORE method. Main Outcome Measures. Apoptosis, proliferation and Fas ligand expression in villous trophoblast. Results. Apoptosis, proliferation and Fas ligand expression were higher in villous trophoblast in HELLP syndrome than in the PE group (p=0.015, p=0.018 and p=0.002, respectively) and the control group (p=0.000, p=0.012 and p=0.049, respectively). Placentae from the PE group had higher levels of apoptosis (p=0.019), lower Fas ligand expression (p=0.029) and no difference in proliferation (p=0.887) compared with the control group. Conclusions. There is an increase in apoptosis, proliferation and Fas ligand expression in placentae from women with HELLP syndrome compared with placentae from PE and normal pregnancies. Our findings indicate the possibility of differential mechanisms behind HELLP syndrome and PE.

Maternal pre-pregnancy underweight and fetal growth in relation to institute of medicine recommendations for gestational weight gain☆

PURPOSE Maternal nutritional status is one of the most important factors of fetal growth and development. Consequently, the currently increasing prevalence of underweight women worldwide has come in the focus of interest of perinatal medicine. The aim of the study was to assess the effect of low pre-pregnancy body mass index (BMI) on fetal growth. MATERIALS AND METHODS Data on 4678 pregnant women and their neonates were retrospectively analyzed. Pre-pregnancy BMI of study women was categorized according to the WHO standards. Fetal growth was assessed by birth weight and birth length, birth weight for gestational age, and ponderal index. RESULTS Study group included 351 (7.6%) women with pregestational BMI<18.5kg/m(2), while all women with pregestational BMI 18.5-25kg/m(2) (n=3688; 78.8%) served as a control group. The mean birth weight and birth length of neonates born to underweight mothers were by 167g and 0.8cm lower in comparison with the neonates born to mothers of normal nutritional status, respectively (P<0.001 both). The prevalence of small for gestational age (SGA) births was twofold that found in the control group of mothers of normal nutritional status (9.7% vs. 4.9%; P<0.001). The inappropriately low gestational weight gain additionally increased the rate of SGA infants in the group of mothers with low pre-pregnancy BMI (21.4% vs. 10.4%; P=0.02). Pre-pregnancy BMI category did not influence neonatal growth symmetry. CONCLUSION Low maternal pregestational BMI is associated with fetal growth assessment. Improvement of the maternal nutritional status before pregnancy can increase the likelihood of perinatal outcome.

Morphological characteristics of placentas associated with idiopathic intrauterine growth retardation: a clinicopathologic study

OBJECTIVES To investigate histopathologic findings, placental diameters and characteristics of syncytial knots in the placentas from idiopathic intrauterine growth retardation (IUGR) pregnancies, and to compare them with a normal birth weight group. STUDY DESIGN Based on strict eligibility criteria, this prospective case-control study included 52 term placentas from idiopathic IUGR pregnancies and 69 term placentas from normal birth weight pregnancies. The study was carried out at the Clinical Hospital Centre, Split, where all placentas were collected and examined. For each placenta, diameters were measured and the following histopathologic findings were recorded: infarction, intervillous thrombosis, abruption, villous branching and maturation, chorioamnionitis, decidual vasculopathy and hemorrhagic endovasculitis for each placenta. In addition we assessed quantitative (number of syncytial knots and number of syncytial nuclei per syncytial knot) and qualitative (density and surface area) characteristics of syncytial knots in each placental sample. Statistical significance was tested using chi(2)-test, Students t-test and Mann-Whitney U-test. Statistical significance was set at P< or =0.05. RESULTS There was no difference in investigated histopathologic findings between idiopathic IUGR placentas and control group placentas. Placental diameters correlated significantly with neonatal birth weight (r=0.64; P<0.01); with higher birth weight there is an increase in placental diameters. Syncytial knots from idiopathic IUGR had significantly smaller surface area (Z=2.637; P=0.008) and higher density (Z=3.225; P=0.001) compared with the control group, while there is no difference in number of syncytial knots per individual villus, total number of syncytial knots in each placenta sample or number of syncytial nuclei per syncytial knot. CONCLUSIONS The investigated histopathologic findings in idiopathic IUGR placentas are incidental, with no higher frequency than in placentas from uncomplicated pregnancies, and should not be considered as possible causative factors for idiopathic IUGR. The demonstrated qualitative changes of syncytial knots in placentas associated with IUGR could represent a compensatory mechanism.

Trophoblast apoptosis in human term placentas from pregnancies complicated with idiopathic intrauterine growth retardation.

Objective. To investigate proliferative, apoptotic, and antiapoptotic activity of placental trophoblast in pregnancies complicated with idiopathic intrauterine growth retardation (IUGR). Methods. Study group included data and placentas from 52 normal singleton term pregnancies with idiopathic IUGR. Records and placentas from 69 singleton pregnancies with normal fetal growth served as a control group. IUGR was defined by birth weight less than 10th percentile of standard values. Children with congenital malformations and those born with the signs of hypoxia, laboratory or clinical signs of preeclampsia or infection, children born to anemic mothers and those born from pregnancies with an increased coagulation system activity were excluded. Results. There was no statistically significant difference in the cytotrophoblast proliferation index value (Z = 0.24; P = 0.553), trophoblast expression of the Bcl-2 antiapoptotic factor (Z = 0.47; P = 0.634), and trophoblast apoptotic index (Z = 0.51; P = 0.613) between the idiopathic IUGR and control group. Conclusion. The proliferative and apoptotic events in the trophoblast of placentas with idiopathic IUGR did not differ from physiologic ones. Study results suggest the IUGR syndrome to have no uniform etiology or even underlying pathophysiology that would determine the possible fetal risk and subsequent long-term consequences for fetal health and life. This imposes the need of a more precise definition and unambiguous distinction between the idiopathic and other forms of IUGR.

Expression of inflammatory cytokines in placentas from pregnancies complicated with preeclampsia and HELLP syndrome

Abstract Objective: To investigate the expression of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-10 (IL-10) in villous trophoblast, syncytial knots and decidua placentas from pregnancies complicated with preeclampsia (PE), Hemolysis, Elevated Liver enzymes and Low Platelet count (HELLP) syndrome and gestational age-matched controls. Methods: Study group included 35 placentas from pregnancies complicated with PE and 35 placentas from pregnancies with HELLP syndrome. Control group included 35 placentas from idiopathic preterm labor. Placentas were matched according to the gestational age. Expression of TNF-α, IL-6 and IL-10 was determined by immunohistochemistry and semi-quantitative HSCORE method in villous trophoblast, syncytial knots and decidua. Non-parametric statistics were used for analyses. Results: There was no difference in the expression of TNF-α, IL-6 and IL-10 in all the studied placental segments between PE, HELLP and gestational age-matched control group. TNF-α (F = 32, 41, p < 0.001), IL-6 (F = 58, 53, p < 0.001) and IL-10 (F = 17, 62, p < 0.001) expression was significantly different in different placental cell types, the highest expression of cytokines was in decidua. Conclusion: There was no difference in cytokine expression in villous trophoblast, syncytial knots and decidua among the studied placental groups. The expression of cytokines was highest in decidua in all the studied placental groups.

Cord blood cortisol level is lower in growth‐restricted newborns

Aim:  To establish the difference in plasma cortisol concentrations between newborns with intrauterine growth‐restricted (IUGR) and appropriate for gestational age (AGA) birthweights.

Low molecular weight heparin treatment and impact of inherited thrombophilia type in pregnancies with previous adverse outcome

Abstract Objective: To assess the impact of low molecular weight heparin (LMWH) treatment in 50 pregnancies of women with inherited thrombophilia and adverse pregnancy outcome (APO) in previous untreated pregnancies. The impact of “Conventional” (FVL, PT, AT, PC, PS) and “Novel” (MTHFR, PAI-1, ACE) thrombophilias on APO was investigated. Methods: The primary outcomes (PO) were: early and late pregnancy loss (EPL, LPL), preterm birth (PTB) or term birth (TB) compared to the last untreated pregnancies of the same women. Secondary outcomes (SO) were APO in LMWH treated and last untreated pregnancies ended with birth. PO and SO were compared in relation to the thrombophilia type. Results: LMWH decreased EPL and LPL rate and improved TB rate compared with last untreated pregnancies (p < 0.001). There were less PTB (p = 0.019) and no cases of intrauterine fetal death (IUFD) (p = 0.0019) in LWMH-treated pregnancies. The division to Conventional and Novel thrombophilias showed: (a) difference between pregnancy losses and birth rate (p = 0.0069) and (b) no difference in the prevalence of APO in untreated pregnancies ended with birth. Conclusions: LMWH treatment improves pregnancy outcome in women with inherited thrombophilia and APO in previous pregnancies. Novel thrombophilias have the equal impact on the pregnancy outcome compared to the Conventional thrombophilias.

Trisomy 1 in an early pregnancy failure

About half of all human conceptions abort spontaneously even before clinical recognition of the pregnancy [Boklage, 1990]. Furthermore, approximately 10–15% of clinically recognized pregnancies end in spontaneous abortion, mostly before the end of the first trimester [NyboAndersen et al., 2000]. Fifty to 70% of these early miscarriages are caused by chromosome abnormalities with the most common abnormality observed being trisomy, accounting for about 60% of all abnormal karyotypes [Philip and Kalousek, 2002]. To date, trisomies of all chromosomes have been observed, and the most rare trisomy is trisomy 1 [Hassold and Jacobs, 1984; Banzai et al., 2004]. Although, cytogenetic analysis of preimplantation embryos, donated from patients undergoing in vitro fertilization (IVF), showed that trisomy 1 does occur [Bahçe et al., 1999], so far only three cases with a full trisomy 1 in a clinically recognized pregnancy have been reported. In these three cases pregnancies were terminated before 9th week of gestation with a blighted ovum [Hanna et al., 1997; Dunn et al., 2001; Banzai et al., 2004]. Herein we present the fourth report of a full trisomy 1 in an early pregnancy failure. This 27-year-old G3PO female was referred for sonographic examination which demonstrated the presence of gestational sac and yolk sac, but the absence of embryonic pole development and fetal heart beat. The diagnosis was blighted ovum and dilatation and curettage (D&C) was performed at 51 days post-last menstrual period (LMP). Products of conception were sent to our laboratory for cytogenetic analysis. Chorionic villi, isolated from products of conception, were cultured and cytogenetic analysis by G-banding showed a 47,XX, þ1,inv(9)(p11q13) karyotype (Fig. 1). Her family history was negative, except for two previous spontaneous abortions. The first, at the age of 25, and the second, at the age of 26, terminated at 9 and 7 weeks of gestation, respectively. Since the couple had infertility, cytogenetic analysis of their blood samples was performed. The father’s karyotype was normal, 46,XY, and the mother’s was 46,XX,inv(9)(p11q13), showing that the variant pericentric inversion of chromosome 9, present in conceptus, was of maternal origin. Autosomal trisomies are found in more than half chromosomally abnormal spontaneous abortions. The most commonly seen is trisomy 16 which accounts for approximately 30% of all trisomies found in miscarriages [Hassold and Jacobs, 1984]. In contrast, trisomy 1 is the rarest, since it has been identified, in only four cases, including our report [Hanna et al., 1997; Dunn et al., 2001; Banzai et al., 2004]. The origin of autosomal trisomies is, in most cases, nondisjunction during maternal meiosis. Although, theoretically all chromosomes should have equal frequency of nondisjunction, Hassold and Jacobs [1984] suggest that the rate of nondisjunction varies among chromosomes and that chromosome 16 has the highest rate among autosomes. Furthermore, chromosomal analysis of human oocytes showed that chromosomes of some groups (A and C) have lower frequency of nondisjunction than expected [Pellestor et al., 2005]. Thus, lower rate of nondisjunction of chromosome 1 could be one of the reasons why trisomy 1 is so infrequent. Although, rarely found in spontaneous abortions, cytogenetic analysis of embryos from patients undergoing IVF showed that trisomy 1 occurs in greater rate in preimplanted embryos [Bahçe et al., 1999]. However, analysis of embryo development showed that chromosomally abnormal embryos, especially those with monosomies and rare trisomies, are