- Email: [email protected]
Oral concurrent session B
Oral Concurrent Session B
Genetics and Prenatal Diagnosis Thursday, February 8, 1996 1:00 p.m.- 3:30 p.m. Kona Ballroom
Moderator: KarinJ. Blakemore, MD Judges:
Mary E. D'Alton, MD W. Patrick Duff, MI) J. Peter Van Dorsten, MD
ABSTRACT NUMBERS: 19-28
310 SPO Abstracts
latm,u~' 1996 Am [ Obslet (.yne(ol
19
THE ANTIOXIDANT LIPOIC ACID - PREVENTS MALFORMATIONS IN OFFSPRING OF DIABETIC RATS. _A Wiznitze#, R. Hershkovitzt, E. Mimca=, M. Mazort, J.R. Leiberman', N. I~lmn =, E.A. Reece. Depts. of OB/GYN & Pediat. Metab. Lab., Soroka Med. err., Ben-Gu~ica Univ., Israel and Dept. of OB/GYN & RS, Tenaple Univ Sch of Med, Phila., PA, USA OBJECTIVE: The ixa-pose of the present investigation was to determine whether LA, an antioxidant, is an effective prophylactic ageat against diabetes-induced embryopathy. STUDY DESIGN: Following conception, lipoic acid, in doses of 10, 30 and 100 mg/kg, was administered intraperitoncnily to 80-day-old Sprague-Dawley rats, five times pet week (pregnancy dayg 2-6). Streptozotocin was injected ca pregnancy day 6 and glucose levels were allowed to remain above 350 mg/dL (normal=150 mg/kg). Four groups of pregnant rats were analyzed for presence of ambryopsthy: In Groups 1 & 2, mothers were under euglyeemic conditions (Gin. 150 mg/dL) with and without LA supplemantatica. Group 3 & 4, were hyperglycemic (Glu. 450 mg/dL) with and without LA supplementation. On pregnancy day 16 conceptuses were examined for size, resorptlc~, anomalies and DNA content. RESULTS: Anomaly rates were significantly high~ in offspring of diabetic rats than among controls (23.5% vs. 3.4%). LA supplementation of 30 mg/kg, resulted in reduction of malformatiotm (23.5% vs. 6.9%) (p <0.001), and fetal loss (17.4% vs. 8.4%) (p <0.005). LA also protected against growth restriction in diabetic ambryos (CRL - 3.6 cmvs. CRL 3.2 cm). The placentas of LA group were essentially protected frown
histologicchanges. CONCLUSIONS: Lipoic acid ccafe~s a protective effect against diabetic embryopathy, fetalloss and embryonic growth restriction.Our data lends further support to the hypothesis that free radicals are causally related to diabetes-associated maldevelopment.
20 L I M B
REDUCTION DEFECTS (LRDs) ARE NOT INCREASED FOLLOWING FIRST TRIMESTER CHORIONIC VILLUS SAMPLING (CVS)R Wapner, L Jackson x, MI Evans, MP Johnson. Divisions o f M F M and Reproductive Genetics, Jefferson Medical College, Philadelphm, PA and Hutzel Hospital, Detroit, MI. O B J E C T I V E : To compare the mcidence of limb reduction defects following chorionic villus sampling to the expected incidence m the general population of 5.6/10,000. STUDY DESIGN: From 8/83-8/95, 19,938 patients have undergone chorionic villus sampling at our insUtutions and have completed theft pregnancies. All patients or thetr physicians were contacted following theft expected date of delivery to determme pregnancy outcome and the presence of congenital abnormalities. The incidence of limb reduction defects was calculated for the enUre patient population and segregated for each week of gestation. RESULTS: There were 6 LRD's not associated with a known genetic syndrome for an incidence of 3/10,000. The gestational age at sampling and the mcidence of limb reduction were: GA # of CVS # of LRD LRD/10,000 births
_
9wks 3,875 1 26
10wks llwks 7,764 5,685 3 2 39 35
12wks 1,380 0 0
>13wks 240 0 0
C O N C L U S I O N : In experienced centers, the incidence of LRDs following CVS ts not increased above the expected for the general population. There is no association between the incidence of LRDs and the gestational age at wMch CVS is performed.
21 THE USE OF SECOND TRIMESTER "GENETIC SONOGRAM" IN GUIDING CLINICAL MANAGEMENT OF PATIENTS AT INCREASED RISK FOR FETAL TRISOMY 21. AM Vintzlleos, WA Campbell, JF Rod=s, ER Guzman, JC Smuhan, DA McLean UMDNJ-RoberI Wood Johnson Medical School/St. Peter's Medical Center, New Brunswick, NJ and UMverstty of Connecticut Health Center, Farmington CT OBJECTIVE' To use ultrasound in the chmca] management of pat=ents at increased risk for fetal tnsomy 21 and also to determtne the efficacy of th~s approach in detecting fetuses with tnsomy 21 STUDY DESIGN. From 1111192to 6/30/95 a second tnmester "genet=c sooogram" was offered to all women with a singleton fetus at increased risk for tnsomy 21 (_>1 274) who either had dechned genetic ammocentests or chose to have a sonogram pnor to dectdtng whether to undergo an ammocentests In addthon to standard fetal b=ometry the following aneuploldy markers were evaluated, struclural anomalies [including face, hands, and cardiac (4-chamber view and outflow tracts)], short femur, short humerus, pyelectasis, nuchal fold thickening, echogenic bowel, choro~d plexus cysts, hypoplasttc mid phalanx of the fifth digit, wide space between the 1st & 2nd toe, and 2-vessel umbilical cord Outcome information included the results of pediatnc assessment and follow-up after birth. RESULTS: A total of 457 patients between 15-23 weeks (mean + SD=19_+1 7) were evaluated, 295 had advanced maternal age (_> 35 years), 121 abnormal serum blochemtstry and 41 had both The majority (391 or 85 5%) had a normal genehc sonogram (absence of any abnormal ultrasound markers), 41 (9%) had 1 marker present and 25 (5 5%) had _>2 markers present Outcome was obtained on 306 patients [the remaintng are ongoing pregnanctes (n=137) or lost to follow-up (n=14)] Eleven of 13 fetuses with tnsomy 21, 1 fetus w~th trisomy 13 and 1 fetus with lnploidy had _>2abnormal ultrasound markers present, 1 fetus wlthtrisomy 21 had 1 abnormal marker and one had a completely normal ultrasound When _>1 abnormal ultrasound markers were present, the sensitivity, specificity, positive and negative pred=cl)ve values for tnsomy 21 were 92 3%, 85.2% 21 8% and 99.6%, respecbvely The overall amnlocentests rate was 13 3% and among cases w=th known outcome was 19 9% CONCLUSION: In expenenced hands, second trimester "genetic sonogram" of high nsk fetuses may result in a htgh detection rate of tnsomy 21 (92 3%) with an amnlocentes=s rate <20%
22 GENETIC
A M N I O C E N T E S I S MAY BE R E A S O N A B L Y AVOIDED IN W O M E N W I T H A B N O R M A L S E R U M SCREENING FOR ANEUPLOIDY BUT NORMAL ULTRASOUND. R. Bahado-Sin~h. A. Tan x, O. Deren x, D. Hunter, J. Copel, J. Mahoneyx Yale University School of Medicine, New Haven CT Norwalk Hospital , Norwalk, CT. OBJECTIVE: To prospectively study the use of ultrasound biometry to refine the risk estimates for both Down syndrome (DS) and any clinically significant chromosome defects (CSCD) in women with abnormal biochemical Triple Screen (TS). STUDY DESIGN' Expected values for humerus length (HL), femur (FL), humerus plus femur (FL+HL), and abdominal circumference (AC) were generated based on blparietal diameter obtained from a separate normal group. Threshold observed + expected (O/E) screening values of each measurement for DS and CSCD screening were determined using receiver operator characteristics curve Using stepwise logistic regression analysis, the optimal screening test, including nuchal thickness (NT) (normal < 6ram) for DS and CSCD detection was determined Tables of blometry - adjusted risk for DS and CSCD for pregnancies with abnormal TS were developed RESULTS' There were 1034 cases with abnormal TS for DS (risk >1/270) or trtsomy 18 (T-18) with 11 cases of DS, 1 T-18 and 17 CSCD. Abnormal NT or O/E HL <0.92 was the most sensitive combination for DS detection. Abnormal NT or t I E FL+HL <0.90 was the most sensitive for CSCD detection. With abnormal blometry or anatomy the DS risk was 8/127 vs 1/753 in normals, OR (95% CI) 50.4 (6.4-90.2), p <0.00001. With abnormal biometry and/or fetal anatomy the rrsk of CSCD was 11/90 vs 6/830 in normals, OR 19.3 (6.4-60.5) p<0.00001 In a pregnancy with 1/270 midtrimester DS risk based on TS, with normal brometry and anatomy the risk falls to 1/2,431. Only one of 609 cases with normal anatomy and btometry and TS risk < 1/50 had DS, (1/609 vs 8/256), p~0.0001. CONCLUSION. Normal anatomy and biometry significantly reduces the risk of aneuploldy m abnormal TS cases and therefore the need for amniocentesis.
Volume 174, Number 1, Part 2 Aln,] Ob~tet G)necol
SPO Abstracts
ELEVATED AMNIOTIC FLUID INTERLEUKIN-6 LEVELS AT GENETIC AMNIOCENTESIS PREDICT SUBSEQUENT PREGNANCY LOSS. KD Wenstromk WW Andrews, T Tamurex, M DuBard x, KE Johnston x, GP Hemstreet. The Unlvarsity of Alabama at Birmingham OBJECTIVE: To determine the proportion of pregnancy loss after genetic amniocentesis that IS related to preexisting subclinical intrauterine inflammation STUDY DESIGN: We accessed our bank of identically stored second trimester amniotic fluid (AF) and maternal serum (MS) samples obtained from all women undergoing genetic amniocentesis at our institution from 1988 to 1995 (n=l 1,971). Interleukm-6 (IL-6) levels were measured by ELISA in samples from every case resulting in spontaneous post procedure loss (excluding aneuploidy and anomalies) within 30 days following the procedure (n=66), and 66 normal controls delivered at term and matched for year of test, gestational age, maternal age, and indication for amntocentesis. RESULTS: Mean MS IL-6 levels were the same in each group (Cases=0 02 - 0.07ng/ml, Controls= 0 06 ± 0 25 ng/ml, p=0.45) Mean AF IL-6 levels were higher in Cases (Cases=4.0 ± 13.1 ng/ml, Controls= 0.5 ± 0.7ng/ml, p = 0.04) The highest AF IL-6 levels tended to occur in Cases with the earliest loss: < 7 days = 8.9 ng/ml, 7-14 days = 4.5 ng/ml, 14-21 days = 3.5 ng/ml, 21-30 days = 1 9 ng/ml. The higher AF mean in the Cases resulted from the inclusion of 8 very high values (>3 S.D. = >2.5 ng/ml). When these samples were excluded, the means and range of values were the same in each group (Cases = 04 ± 04 ng/ml, Controls = 0.5 ± 0.Tng/ml, p = 0.58). Twelve per cent (8/66) of the Cases and 3% (2/66) of the Controls had AF IL-6 values > 2.5 ng/ml (p = 0.048, Odds RaUo = 4.1,95% C I = 1 0 - 31 2). Although the overall correlation between MS IL-6 and AF IL-6 levels was good (r = 0.50, p = 0 0015), only 1 of the 8 Cases with high AF IL-6 would have been identified by a MS IL-6 > 3 S.D. (:> 0.8 ng/ml). CONCLUSIONS: Analysis of our complete, unselected group of post-amniocentesis pregnancy losses indicates that up to 12% may result from preexisting subclinical intrauterine inflammation. This ,nflammation is Iocahzed and not reflected by high maternal serum IL6 levels; such cases cannot be identified by maternal serum testing pnor to the procedure.
23
ENDOSCOPIC TRACHEAL PLUGGING USING AN INFLATABLE BALLOOh~ IN THE FETAL LAMB. I ~ JA*, Ewerd VA, Van Ballaer PP, Verbeken EA. Vandenberghe K, Breserts IA, Van Aesche FA, Lerut T Centre for Surgical Technologies, KU Lenven, Depts Ob/Gyn & Pathology. Univ HospRals "Gasthuisberg", 3000 Leuven, Belgium OBJECTIVE. To evaluate the feasibility and pulmonary effects of intra-tracheal obstructlon by a detachable balloon, andoscopically pesamned by fetaltracheoscopy STUDY DESIGN: A case-controlled surgical trial was performed m 13 time-dated pregnant ewes. 15 fetuses were subjected to tracheoscoplc balloon obstmchon during a progresswely longer period (range 2-18 days). In case of multiple pregnancies, other fetuses (n=10) were used as negative controls. A 1 2 mm mmmcope with a double lumen shent, allowing low flow tmgatinn and a coaxial catheter loaded with a detachable balloon, was used for tracheoseopic plugging. The fust nine fetuses (Geetatlonal Age (GA) 95-120 days; term=145 days) were used as a pilot group to develop the techmque and to assess the obstructive performance oftlus plug A second group ofs,x mid-trimester fetuses (GA 90-99 days) was allowed a longer follow-up (range 14-18 days) to assess pulmonary effects, using lung-to-body weight ratm (LBWR) and morphometnc terminal brochml density (MTBD). Feasibility of the tochmque was assessed in all animals using operation t~me, intra-operative complieaUons, and tracheal obstructionas outcome parameters RESULTS: The first balloon failed to remain inflated due to an evitable valve failure In the other 14 animals, the trachea was successfully obstructed till dehvery Tracheoscop~c mampulation time ranged from 3 to 14 minutes. One mtra-operatwe death occurred, but the contralateral control fetus also died during the procedure In the second group of 6 treated mid.tranester fetuses mean LBWR was 0.060 + 0 01 (0.051-0 075) whtle m controls 0.031 + 0.01 (range:0.017-0.039, P<00005). MTBD was 065-2059, as compared to 1.30 4-0.80 for controls. CONCLUSION" Using a sunple and fast techmque of fetoscop~e tracheoscopy, the fetal trachea was successfullyobstructed w~th an inflatable balloon Pulmonary hyperplasm, of potential value when treating Congenital Diaphragmatic Herraa, was demonstrated
25
24
INTRA-AMNIOTIC PRESSURE REDUCTION IN TWIN-TO-TWIN TRANSFUSION SYNDROME Garry D~Lysikiewicz A, Mays J,~Tejam N New York M¢&cal College, Valhalla, NY OBJECTIVE: Serial ammooentcsls has been performed in the treatment of twin-to-twin transfusmn syndrome (TTTS) Reduction m intra-ammotlc pressure (IAP) has been proposed as the reason for favorable outcome although th~s has never been demonstrated Our purpose was to evaluate IAP before and after docompressmn ammocentests m TITS STUDY DESIGN: Docompressmn ammocentesis for hydrammos was performed m the largest amniotlc fired (AF) pocket on 9 occasmns m 3 TITS patients The IAP was determined with a water manometer before and after drmnage of AF The manometer was referenced to the top of the maternal abdomen AF was removed untd the single pocket measured g-10 cm ver'acany The procedure was repeated every 7-10 days untd AF mdexes equalLzed m the two sacs Patients were followed through dehvery The IAP was also determined in the same manner in 5 singleton gestations and measurements compared with reported values Results were analyzed using t-tests RESULTS: The mean IAP in normal sinsleton gestatmns was 8 01cm H20 +_3 8 The mean initial IAP in the TTTS patients was 19 2 cm H20 +-5 67 in the hydramniouc sac AF pressure fell significantly (mean pressure change -5 02 cm H20, 95% CI - 2 44 to- 7 62, p = 001) m g of ~..~.~.~ ..b ~..~ .d ~ ~ . ~ the 9 measurements The moan ending "i " ~ " ~"~'a''''T pressure was 14 2 cm H20 +5 08 which " ] was not &fferent from normal singleton ~ I _ ~ m I measurements. The volume of AF removed •[ ~ ~ IBm. ,~ I • es equalg ed • "L..J.~" ....... ranged from 500 to 1800 cc AF index . . . . . . . . .7~."=" . . . . .J?J: ......... i m all cases including 1 "stuck" twin. 6 neonates '.... vathameanblrthweightof1663gm+802for ','. : . " 2 . , ~ ~-~, :. .~ .~' twln A and 1033 gm +-255 8 for twin B were ~,m~m~J,0 delivered at an average gestatmnal age 0f32 weeks +1 7 CONCLUSION: RepeUbve docompressmn ammooentesls for treatment of TTTS sigmficantly reduces the IAP The ending pressure may be used to guide AF removal volume•
26 ALTERED EXPRESSION OF PLA2 GENE IMPLICATED IN MOLECULAR MECHANISM OF DIABETES-INDUCED NEURAL TUBE DEFECTS (NTIh): A N E W R E L E V A T I O N . E.A. Reece. YK. Wu', A. Ait-Allah ~, W. Salamehz. Deperlmmt of OB/GYN & RS, Temple Uulv Sch of Med, Phila., PA OBJECTIVE: Hyperglycemia has been shown to induce NTDs and cytm~hitectural changes in the embryonic neuroepitbelium and yolk sac. This was shown to be causally related to an arachi&mic acid (AA) deficiency state. The object oi' our studies was to detexmine whether the decrease in AA resulted from altered PLA~ gone expression. STUDY DESIGN: 80-dey old Sprague Dawley rats wea-e mated, and following couceptiou were randomly assigned to s diabetes and a control group. Diabetes was induced by streptozotocin (65 mg/kg). Sertma levels of arachidonic acid (AA) wexe obtained. Animals were sacrificed on Day 12; conceptuscs (embryos and yolk sacs) were eJamined for malformations; mRNAs wexe extracted and Ncrdicrn blot analysis performed using radiolabelled PLA2 probe. After standard processing for in situ hybridization, yolk sac and embryo sections were hybridized with PLA2-digoxigenin probe, Signal was detected by alkaline phosphatasetagged antidoxigenin antibody. RESULTS: As we have previously reported, diabetic rats had a significantly higher malfcrmatious than contn31s (22.4% vs. 3.4%; p <0.001); lower AA levels (17.83±5.84 pg/ml vs. 14.18±2.58 b~g/ml; p <0.001); increased absorption and decreased conceptus size in coml~rison to controls. In situ hybridization revealed dislribution of PLA2 mRNA in neuroepithelium and yolk sac of non-diabetic coulTols, but marl~dly reduced gene expression among diabetics, coincident with reduced serum AA levels and embryopathy. CONCLUSIONS: We have demonstrated for the first time that hyperglycemia Induces a reduced expression of PLAz gape a~ivity, which results In an AA deficiency state and embryopathy. This novel finding advances a molecular basis for DM-induced NTDs.
31
312
SPO Abstracts
,lanuar~ 1996 A m , ] Obstet Gynecol
27
ENGRAFFMENT FOLLOWING IN U'lEtgO BONE MARROW TRANSPLANTATION FOR GLOBOID CELL LEUKODYSTROPHY. K. Biakemore. B. Bsmbach, z H. M o s e r / V . Corson/C. O r i f f m / S . Noga? E. Perhnan," D. Weager? R. Znckerman,~ A. Khouzami," R. Jones. ~ Dapts. Gyn/Ob, Neurol, Neuroganetks, Pathol, Anesthesinl, Oncology, Johns Hopkins Univ. Sch. of Med. & Kennedy Krieger Inst., Baltimore, MD and Dept. Pedlatr, Thomas Jefferson Univ., Philadelphia, PA. OBJECTIVE: To study whether signifw.ant host engraftment c4m occur following/n uSero bone marrow ummplantation (BMT) during the f~rst trimeger using CD34+-solection. STUDY DESIGN: Following genetic counseling and informed consent under an IRB/FDA-approved study protocol,/n ~tero ~ w u performed at 13 3/7 weeks of gestation on n fetus with globoid cell Icukodystrophy diagnosed by chorionic villus sampling. 2.4 x 10a° nucleated bone marrow cells were harvested from the father, the mononucleac cells concentrated, and the CD34 + ceils isolated in n final volume of 0.8 ml following positive selection with the CellPrn Caprete SC Stem Cell Concentrator. Flow cytometry revealed 91% purity for CD34" cells (1.4 x 10') with 5% (7 x 10') T cells; thea,e were transfnsod into the fetal peritoneal cavity under ultrasonnd guidance using n 22gauge spinal needle. RESULTS: Sonograms at O, 4 and 14 hours, and at 1 and 5 weckJ after BMT showed normal fetal activity, heart rate, and growth. At 20 weskl, fetal demise was diagnosed. Following induction of labor, aa autopsy and engrnflmeat stndies were performed. Histology revealed exte~ive and widespread extramedullary hematopoiesis. Hemorrhage was present in the lungs and liver. DNA-RFLP stndies showed significant percantagas of donor ceils in the liver, spleen, and Ikin ( - 9 5 % , 50%, and 9% paternal in origin, respectively). CONCLUSION: Onr protocol is the first to enable substantiul engraftment of a fetus without n primary immunodofiulency disorder following/n utero BM'r. The enhanced donor angrnflment likely resulted from the early gegationel age and CD34+-solactlon allowing greater &-JI numbers in a very small volume. A 10-fuld decrease in CD34 + cells is planned for future studies. In utero BMT hag the potential to correct a variety of disorders dingnosablc prenatally.
28
FETAL BONE MARROW ORGAN CULTURE S.Cntrmel." C.O'Doanell/ C. Ulrich? T.Crombleholme/ The Fetid Treatment Program, New England Medical Center/Tufts University School of Medicine, Boston, MA OBJECTIVE: We developed an in vitro model of fetal bone marrow organ culture (BMOC) in order to study the normal interaction of the stem cell and the developing stroma as well as provide an in vitro model of tn utero ~ cell transplantation. STUDY DESIGN: The fetuses of pregnant rabbits were harvested on day 27 of gestation and the long bones were dissected under sterile conditions. Thin sections of fetal bone were placed into 30mm culture wells on filter paper scaffolding. Culture~ were fed five times a week with IMDM, penicillin/ straptemyuln (100 units ench/ml), 20% fetal calf serum, and hydrocortisene ( I x 10"~VI). Each BMOC was cultured with either Interleukin-3 (IL-3, 25 ng/ml) and Granulocyte-Macrophnge-ColonyStimulating Factor (GM-CSF, 25ng/ml) or Interleukin-6 (IL-6, lng/ml). Progenitor assays were performed on the effluent after 14 days and the presence of myelold, erythrold, and lymphoid precursors confirmed by histologlc examination. RESULTS: Fetal BMOC successfully supported fetal bone marrow cells and stroma in culture out to 9 weeks. The vast majority of colonies were of the Colony Forming Unit, Granulecyte-Macrophagn (CFU-GM) type. The maximal number of colonies were seen between five and seven weeks of culture. This held true for BMOC cultured with IL-3 and GM-CSF as well as BMOC cultured with IL-6. CONCLUSIONS: BMOC is a viable/n vitro model for the study of NSCstromul interactions. IMDM with growth factor edditivas was found to support BMOC growth for at least 9 weeks. This/n vitro model will provide n useful tool for the study of stem cell-stromel interactions, regulatory mechanisms controlling hematopoiesis and engraftment, and in utero HSC transplantation.
Genetics and Prenatal Diagnosis Thursday, February 8, 1996 1:00 p.m.- 3:30 p.m. Kona Ballroom
Moderator: KarinJ. Blakemore, MD Judges:
Mary E. D'Alton, MD W. Patrick Duff, MI) J. Peter Van Dorsten, MD
ABSTRACT NUMBERS: 19-28
310 SPO Abstracts
latm,u~' 1996 Am [ Obslet (.yne(ol
19
THE ANTIOXIDANT LIPOIC ACID - PREVENTS MALFORMATIONS IN OFFSPRING OF DIABETIC RATS. _A Wiznitze#, R. Hershkovitzt, E. Mimca=, M. Mazort, J.R. Leiberman', N. I~lmn =, E.A. Reece. Depts. of OB/GYN & Pediat. Metab. Lab., Soroka Med. err., Ben-Gu~ica Univ., Israel and Dept. of OB/GYN & RS, Tenaple Univ Sch of Med, Phila., PA, USA OBJECTIVE: The ixa-pose of the present investigation was to determine whether LA, an antioxidant, is an effective prophylactic ageat against diabetes-induced embryopathy. STUDY DESIGN: Following conception, lipoic acid, in doses of 10, 30 and 100 mg/kg, was administered intraperitoncnily to 80-day-old Sprague-Dawley rats, five times pet week (pregnancy dayg 2-6). Streptozotocin was injected ca pregnancy day 6 and glucose levels were allowed to remain above 350 mg/dL (normal=150 mg/kg). Four groups of pregnant rats were analyzed for presence of ambryopsthy: In Groups 1 & 2, mothers were under euglyeemic conditions (Gin. 150 mg/dL) with and without LA supplemantatica. Group 3 & 4, were hyperglycemic (Glu. 450 mg/dL) with and without LA supplementation. On pregnancy day 16 conceptuses were examined for size, resorptlc~, anomalies and DNA content. RESULTS: Anomaly rates were significantly high~ in offspring of diabetic rats than among controls (23.5% vs. 3.4%). LA supplementation of 30 mg/kg, resulted in reduction of malformatiotm (23.5% vs. 6.9%) (p <0.001), and fetal loss (17.4% vs. 8.4%) (p <0.005). LA also protected against growth restriction in diabetic ambryos (CRL - 3.6 cmvs. CRL 3.2 cm). The placentas of LA group were essentially protected frown
histologicchanges. CONCLUSIONS: Lipoic acid ccafe~s a protective effect against diabetic embryopathy, fetalloss and embryonic growth restriction.Our data lends further support to the hypothesis that free radicals are causally related to diabetes-associated maldevelopment.
20 L I M B
REDUCTION DEFECTS (LRDs) ARE NOT INCREASED FOLLOWING FIRST TRIMESTER CHORIONIC VILLUS SAMPLING (CVS)R Wapner, L Jackson x, MI Evans, MP Johnson. Divisions o f M F M and Reproductive Genetics, Jefferson Medical College, Philadelphm, PA and Hutzel Hospital, Detroit, MI. O B J E C T I V E : To compare the mcidence of limb reduction defects following chorionic villus sampling to the expected incidence m the general population of 5.6/10,000. STUDY DESIGN: From 8/83-8/95, 19,938 patients have undergone chorionic villus sampling at our insUtutions and have completed theft pregnancies. All patients or thetr physicians were contacted following theft expected date of delivery to determme pregnancy outcome and the presence of congenital abnormalities. The incidence of limb reduction defects was calculated for the enUre patient population and segregated for each week of gestation. RESULTS: There were 6 LRD's not associated with a known genetic syndrome for an incidence of 3/10,000. The gestational age at sampling and the mcidence of limb reduction were: GA # of CVS # of LRD LRD/10,000 births
_
9wks 3,875 1 26
10wks llwks 7,764 5,685 3 2 39 35
12wks 1,380 0 0
>13wks 240 0 0
C O N C L U S I O N : In experienced centers, the incidence of LRDs following CVS ts not increased above the expected for the general population. There is no association between the incidence of LRDs and the gestational age at wMch CVS is performed.
21 THE USE OF SECOND TRIMESTER "GENETIC SONOGRAM" IN GUIDING CLINICAL MANAGEMENT OF PATIENTS AT INCREASED RISK FOR FETAL TRISOMY 21. AM Vintzlleos, WA Campbell, JF Rod=s, ER Guzman, JC Smuhan, DA McLean UMDNJ-RoberI Wood Johnson Medical School/St. Peter's Medical Center, New Brunswick, NJ and UMverstty of Connecticut Health Center, Farmington CT OBJECTIVE' To use ultrasound in the chmca] management of pat=ents at increased risk for fetal tnsomy 21 and also to determtne the efficacy of th~s approach in detecting fetuses with tnsomy 21 STUDY DESIGN. From 1111192to 6/30/95 a second tnmester "genet=c sooogram" was offered to all women with a singleton fetus at increased risk for tnsomy 21 (_>1 274) who either had dechned genetic ammocentests or chose to have a sonogram pnor to dectdtng whether to undergo an ammocentests In addthon to standard fetal b=ometry the following aneuploldy markers were evaluated, struclural anomalies [including face, hands, and cardiac (4-chamber view and outflow tracts)], short femur, short humerus, pyelectasis, nuchal fold thickening, echogenic bowel, choro~d plexus cysts, hypoplasttc mid phalanx of the fifth digit, wide space between the 1st & 2nd toe, and 2-vessel umbilical cord Outcome information included the results of pediatnc assessment and follow-up after birth. RESULTS: A total of 457 patients between 15-23 weeks (mean + SD=19_+1 7) were evaluated, 295 had advanced maternal age (_> 35 years), 121 abnormal serum blochemtstry and 41 had both The majority (391 or 85 5%) had a normal genehc sonogram (absence of any abnormal ultrasound markers), 41 (9%) had 1 marker present and 25 (5 5%) had _>2 markers present Outcome was obtained on 306 patients [the remaintng are ongoing pregnanctes (n=137) or lost to follow-up (n=14)] Eleven of 13 fetuses with tnsomy 21, 1 fetus w~th trisomy 13 and 1 fetus with lnploidy had _>2abnormal ultrasound markers present, 1 fetus wlthtrisomy 21 had 1 abnormal marker and one had a completely normal ultrasound When _>1 abnormal ultrasound markers were present, the sensitivity, specificity, positive and negative pred=cl)ve values for tnsomy 21 were 92 3%, 85.2% 21 8% and 99.6%, respecbvely The overall amnlocentests rate was 13 3% and among cases w=th known outcome was 19 9% CONCLUSION: In expenenced hands, second trimester "genetic sonogram" of high nsk fetuses may result in a htgh detection rate of tnsomy 21 (92 3%) with an amnlocentes=s rate <20%
22 GENETIC
A M N I O C E N T E S I S MAY BE R E A S O N A B L Y AVOIDED IN W O M E N W I T H A B N O R M A L S E R U M SCREENING FOR ANEUPLOIDY BUT NORMAL ULTRASOUND. R. Bahado-Sin~h. A. Tan x, O. Deren x, D. Hunter, J. Copel, J. Mahoneyx Yale University School of Medicine, New Haven CT Norwalk Hospital , Norwalk, CT. OBJECTIVE: To prospectively study the use of ultrasound biometry to refine the risk estimates for both Down syndrome (DS) and any clinically significant chromosome defects (CSCD) in women with abnormal biochemical Triple Screen (TS). STUDY DESIGN' Expected values for humerus length (HL), femur (FL), humerus plus femur (FL+HL), and abdominal circumference (AC) were generated based on blparietal diameter obtained from a separate normal group. Threshold observed + expected (O/E) screening values of each measurement for DS and CSCD screening were determined using receiver operator characteristics curve Using stepwise logistic regression analysis, the optimal screening test, including nuchal thickness (NT) (normal < 6ram) for DS and CSCD detection was determined Tables of blometry - adjusted risk for DS and CSCD for pregnancies with abnormal TS were developed RESULTS' There were 1034 cases with abnormal TS for DS (risk >1/270) or trtsomy 18 (T-18) with 11 cases of DS, 1 T-18 and 17 CSCD. Abnormal NT or O/E HL <0.92 was the most sensitive combination for DS detection. Abnormal NT or t I E FL+HL <0.90 was the most sensitive for CSCD detection. With abnormal blometry or anatomy the DS risk was 8/127 vs 1/753 in normals, OR (95% CI) 50.4 (6.4-90.2), p <0.00001. With abnormal biometry and/or fetal anatomy the rrsk of CSCD was 11/90 vs 6/830 in normals, OR 19.3 (6.4-60.5) p<0.00001 In a pregnancy with 1/270 midtrimester DS risk based on TS, with normal brometry and anatomy the risk falls to 1/2,431. Only one of 609 cases with normal anatomy and btometry and TS risk < 1/50 had DS, (1/609 vs 8/256), p~0.0001. CONCLUSION. Normal anatomy and biometry significantly reduces the risk of aneuploldy m abnormal TS cases and therefore the need for amniocentesis.
Volume 174, Number 1, Part 2 Aln,] Ob~tet G)necol
SPO Abstracts
ELEVATED AMNIOTIC FLUID INTERLEUKIN-6 LEVELS AT GENETIC AMNIOCENTESIS PREDICT SUBSEQUENT PREGNANCY LOSS. KD Wenstromk WW Andrews, T Tamurex, M DuBard x, KE Johnston x, GP Hemstreet. The Unlvarsity of Alabama at Birmingham OBJECTIVE: To determine the proportion of pregnancy loss after genetic amniocentesis that IS related to preexisting subclinical intrauterine inflammation STUDY DESIGN: We accessed our bank of identically stored second trimester amniotic fluid (AF) and maternal serum (MS) samples obtained from all women undergoing genetic amniocentesis at our institution from 1988 to 1995 (n=l 1,971). Interleukm-6 (IL-6) levels were measured by ELISA in samples from every case resulting in spontaneous post procedure loss (excluding aneuploidy and anomalies) within 30 days following the procedure (n=66), and 66 normal controls delivered at term and matched for year of test, gestational age, maternal age, and indication for amntocentesis. RESULTS: Mean MS IL-6 levels were the same in each group (Cases=0 02 - 0.07ng/ml, Controls= 0 06 ± 0 25 ng/ml, p=0.45) Mean AF IL-6 levels were higher in Cases (Cases=4.0 ± 13.1 ng/ml, Controls= 0.5 ± 0.7ng/ml, p = 0.04) The highest AF IL-6 levels tended to occur in Cases with the earliest loss: < 7 days = 8.9 ng/ml, 7-14 days = 4.5 ng/ml, 14-21 days = 3.5 ng/ml, 21-30 days = 1 9 ng/ml. The higher AF mean in the Cases resulted from the inclusion of 8 very high values (>3 S.D. = >2.5 ng/ml). When these samples were excluded, the means and range of values were the same in each group (Cases = 04 ± 04 ng/ml, Controls = 0.5 ± 0.Tng/ml, p = 0.58). Twelve per cent (8/66) of the Cases and 3% (2/66) of the Controls had AF IL-6 values > 2.5 ng/ml (p = 0.048, Odds RaUo = 4.1,95% C I = 1 0 - 31 2). Although the overall correlation between MS IL-6 and AF IL-6 levels was good (r = 0.50, p = 0 0015), only 1 of the 8 Cases with high AF IL-6 would have been identified by a MS IL-6 > 3 S.D. (:> 0.8 ng/ml). CONCLUSIONS: Analysis of our complete, unselected group of post-amniocentesis pregnancy losses indicates that up to 12% may result from preexisting subclinical intrauterine inflammation. This ,nflammation is Iocahzed and not reflected by high maternal serum IL6 levels; such cases cannot be identified by maternal serum testing pnor to the procedure.
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ENDOSCOPIC TRACHEAL PLUGGING USING AN INFLATABLE BALLOOh~ IN THE FETAL LAMB. I ~ JA*, Ewerd VA, Van Ballaer PP, Verbeken EA. Vandenberghe K, Breserts IA, Van Aesche FA, Lerut T Centre for Surgical Technologies, KU Lenven, Depts Ob/Gyn & Pathology. Univ HospRals "Gasthuisberg", 3000 Leuven, Belgium OBJECTIVE. To evaluate the feasibility and pulmonary effects of intra-tracheal obstructlon by a detachable balloon, andoscopically pesamned by fetaltracheoscopy STUDY DESIGN: A case-controlled surgical trial was performed m 13 time-dated pregnant ewes. 15 fetuses were subjected to tracheoscoplc balloon obstmchon during a progresswely longer period (range 2-18 days). In case of multiple pregnancies, other fetuses (n=10) were used as negative controls. A 1 2 mm mmmcope with a double lumen shent, allowing low flow tmgatinn and a coaxial catheter loaded with a detachable balloon, was used for tracheoseopic plugging. The fust nine fetuses (Geetatlonal Age (GA) 95-120 days; term=145 days) were used as a pilot group to develop the techmque and to assess the obstructive performance oftlus plug A second group ofs,x mid-trimester fetuses (GA 90-99 days) was allowed a longer follow-up (range 14-18 days) to assess pulmonary effects, using lung-to-body weight ratm (LBWR) and morphometnc terminal brochml density (MTBD). Feasibility of the tochmque was assessed in all animals using operation t~me, intra-operative complieaUons, and tracheal obstructionas outcome parameters RESULTS: The first balloon failed to remain inflated due to an evitable valve failure In the other 14 animals, the trachea was successfully obstructed till dehvery Tracheoscop~c mampulation time ranged from 3 to 14 minutes. One mtra-operatwe death occurred, but the contralateral control fetus also died during the procedure In the second group of 6 treated mid.tranester fetuses mean LBWR was 0.060 + 0 01 (0.051-0 075) whtle m controls 0.031 + 0.01 (range:0.017-0.039, P<00005). MTBD was 065-2059, as compared to 1.30 4-0.80 for controls. CONCLUSION" Using a sunple and fast techmque of fetoscop~e tracheoscopy, the fetal trachea was successfullyobstructed w~th an inflatable balloon Pulmonary hyperplasm, of potential value when treating Congenital Diaphragmatic Herraa, was demonstrated
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INTRA-AMNIOTIC PRESSURE REDUCTION IN TWIN-TO-TWIN TRANSFUSION SYNDROME Garry D~Lysikiewicz A, Mays J,~Tejam N New York M¢&cal College, Valhalla, NY OBJECTIVE: Serial ammooentcsls has been performed in the treatment of twin-to-twin transfusmn syndrome (TTTS) Reduction m intra-ammotlc pressure (IAP) has been proposed as the reason for favorable outcome although th~s has never been demonstrated Our purpose was to evaluate IAP before and after docompressmn ammocentests m TITS STUDY DESIGN: Docompressmn ammocentesis for hydrammos was performed m the largest amniotlc fired (AF) pocket on 9 occasmns m 3 TITS patients The IAP was determined with a water manometer before and after drmnage of AF The manometer was referenced to the top of the maternal abdomen AF was removed untd the single pocket measured g-10 cm ver'acany The procedure was repeated every 7-10 days untd AF mdexes equalLzed m the two sacs Patients were followed through dehvery The IAP was also determined in the same manner in 5 singleton gestations and measurements compared with reported values Results were analyzed using t-tests RESULTS: The mean IAP in normal sinsleton gestatmns was 8 01cm H20 +_3 8 The mean initial IAP in the TTTS patients was 19 2 cm H20 +-5 67 in the hydramniouc sac AF pressure fell significantly (mean pressure change -5 02 cm H20, 95% CI - 2 44 to- 7 62, p = 001) m g of ~..~.~.~ ..b ~..~ .d ~ ~ . ~ the 9 measurements The moan ending "i " ~ " ~"~'a''''T pressure was 14 2 cm H20 +5 08 which " ] was not &fferent from normal singleton ~ I _ ~ m I measurements. The volume of AF removed •[ ~ ~ IBm. ,~ I • es equalg ed • "L..J.~" ....... ranged from 500 to 1800 cc AF index . . . . . . . . .7~."=" . . . . .J?J: ......... i m all cases including 1 "stuck" twin. 6 neonates '.... vathameanblrthweightof1663gm+802for ','. : . " 2 . , ~ ~-~, :. .~ .~' twln A and 1033 gm +-255 8 for twin B were ~,m~m~J,0 delivered at an average gestatmnal age 0f32 weeks +1 7 CONCLUSION: RepeUbve docompressmn ammooentesls for treatment of TTTS sigmficantly reduces the IAP The ending pressure may be used to guide AF removal volume•
26 ALTERED EXPRESSION OF PLA2 GENE IMPLICATED IN MOLECULAR MECHANISM OF DIABETES-INDUCED NEURAL TUBE DEFECTS (NTIh): A N E W R E L E V A T I O N . E.A. Reece. YK. Wu', A. Ait-Allah ~, W. Salamehz. Deperlmmt of OB/GYN & RS, Temple Uulv Sch of Med, Phila., PA OBJECTIVE: Hyperglycemia has been shown to induce NTDs and cytm~hitectural changes in the embryonic neuroepitbelium and yolk sac. This was shown to be causally related to an arachi&mic acid (AA) deficiency state. The object oi' our studies was to detexmine whether the decrease in AA resulted from altered PLA~ gone expression. STUDY DESIGN: 80-dey old Sprague Dawley rats wea-e mated, and following couceptiou were randomly assigned to s diabetes and a control group. Diabetes was induced by streptozotocin (65 mg/kg). Sertma levels of arachidonic acid (AA) wexe obtained. Animals were sacrificed on Day 12; conceptuscs (embryos and yolk sacs) were eJamined for malformations; mRNAs wexe extracted and Ncrdicrn blot analysis performed using radiolabelled PLA2 probe. After standard processing for in situ hybridization, yolk sac and embryo sections were hybridized with PLA2-digoxigenin probe, Signal was detected by alkaline phosphatasetagged antidoxigenin antibody. RESULTS: As we have previously reported, diabetic rats had a significantly higher malfcrmatious than contn31s (22.4% vs. 3.4%; p <0.001); lower AA levels (17.83±5.84 pg/ml vs. 14.18±2.58 b~g/ml; p <0.001); increased absorption and decreased conceptus size in coml~rison to controls. In situ hybridization revealed dislribution of PLA2 mRNA in neuroepithelium and yolk sac of non-diabetic coulTols, but marl~dly reduced gene expression among diabetics, coincident with reduced serum AA levels and embryopathy. CONCLUSIONS: We have demonstrated for the first time that hyperglycemia Induces a reduced expression of PLAz gape a~ivity, which results In an AA deficiency state and embryopathy. This novel finding advances a molecular basis for DM-induced NTDs.
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312
SPO Abstracts
,lanuar~ 1996 A m , ] Obstet Gynecol
27
ENGRAFFMENT FOLLOWING IN U'lEtgO BONE MARROW TRANSPLANTATION FOR GLOBOID CELL LEUKODYSTROPHY. K. Biakemore. B. Bsmbach, z H. M o s e r / V . Corson/C. O r i f f m / S . Noga? E. Perhnan," D. Weager? R. Znckerman,~ A. Khouzami," R. Jones. ~ Dapts. Gyn/Ob, Neurol, Neuroganetks, Pathol, Anesthesinl, Oncology, Johns Hopkins Univ. Sch. of Med. & Kennedy Krieger Inst., Baltimore, MD and Dept. Pedlatr, Thomas Jefferson Univ., Philadelphia, PA. OBJECTIVE: To study whether signifw.ant host engraftment c4m occur following/n uSero bone marrow ummplantation (BMT) during the f~rst trimeger using CD34+-solection. STUDY DESIGN: Following genetic counseling and informed consent under an IRB/FDA-approved study protocol,/n ~tero ~ w u performed at 13 3/7 weeks of gestation on n fetus with globoid cell Icukodystrophy diagnosed by chorionic villus sampling. 2.4 x 10a° nucleated bone marrow cells were harvested from the father, the mononucleac cells concentrated, and the CD34 + ceils isolated in n final volume of 0.8 ml following positive selection with the CellPrn Caprete SC Stem Cell Concentrator. Flow cytometry revealed 91% purity for CD34" cells (1.4 x 10') with 5% (7 x 10') T cells; thea,e were transfnsod into the fetal peritoneal cavity under ultrasonnd guidance using n 22gauge spinal needle. RESULTS: Sonograms at O, 4 and 14 hours, and at 1 and 5 weckJ after BMT showed normal fetal activity, heart rate, and growth. At 20 weskl, fetal demise was diagnosed. Following induction of labor, aa autopsy and engrnflmeat stndies were performed. Histology revealed exte~ive and widespread extramedullary hematopoiesis. Hemorrhage was present in the lungs and liver. DNA-RFLP stndies showed significant percantagas of donor ceils in the liver, spleen, and Ikin ( - 9 5 % , 50%, and 9% paternal in origin, respectively). CONCLUSION: Onr protocol is the first to enable substantiul engraftment of a fetus without n primary immunodofiulency disorder following/n utero BM'r. The enhanced donor angrnflment likely resulted from the early gegationel age and CD34+-solactlon allowing greater &-JI numbers in a very small volume. A 10-fuld decrease in CD34 + cells is planned for future studies. In utero BMT hag the potential to correct a variety of disorders dingnosablc prenatally.
28
FETAL BONE MARROW ORGAN CULTURE S.Cntrmel." C.O'Doanell/ C. Ulrich? T.Crombleholme/ The Fetid Treatment Program, New England Medical Center/Tufts University School of Medicine, Boston, MA OBJECTIVE: We developed an in vitro model of fetal bone marrow organ culture (BMOC) in order to study the normal interaction of the stem cell and the developing stroma as well as provide an in vitro model of tn utero ~ cell transplantation. STUDY DESIGN: The fetuses of pregnant rabbits were harvested on day 27 of gestation and the long bones were dissected under sterile conditions. Thin sections of fetal bone were placed into 30mm culture wells on filter paper scaffolding. Culture~ were fed five times a week with IMDM, penicillin/ straptemyuln (100 units ench/ml), 20% fetal calf serum, and hydrocortisene ( I x 10"~VI). Each BMOC was cultured with either Interleukin-3 (IL-3, 25 ng/ml) and Granulocyte-Macrophnge-ColonyStimulating Factor (GM-CSF, 25ng/ml) or Interleukin-6 (IL-6, lng/ml). Progenitor assays were performed on the effluent after 14 days and the presence of myelold, erythrold, and lymphoid precursors confirmed by histologlc examination. RESULTS: Fetal BMOC successfully supported fetal bone marrow cells and stroma in culture out to 9 weeks. The vast majority of colonies were of the Colony Forming Unit, Granulecyte-Macrophagn (CFU-GM) type. The maximal number of colonies were seen between five and seven weeks of culture. This held true for BMOC cultured with IL-3 and GM-CSF as well as BMOC cultured with IL-6. CONCLUSIONS: BMOC is a viable/n vitro model for the study of NSCstromul interactions. IMDM with growth factor edditivas was found to support BMOC growth for at least 9 weeks. This/n vitro model will provide n useful tool for the study of stem cell-stromel interactions, regulatory mechanisms controlling hematopoiesis and engraftment, and in utero HSC transplantation.