University Department of Obstetrics and Gynaecology, Liverpool Women's Hospital, UK
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Abstract |
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Key words: adverse pregnancy outcome/placental thrombosis/thrombophilia
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Introduction |
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The aim of the present study was to provide additional data on the relationship between placental changes and thrombophilia status in women with serious pregnancy complications.
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Materials and methods |
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Cases with fetal congenital malformation, abnormal karyotype or known maternal history of deep venous thrombosis or pulmonary embolism were excluded.
Thrombophilia screen
All blood tests were done at least 10 weeks after delivery and included assays of antithrombin III, protein C activity, antigen protein S free and total, activated protein C resistance (APCR), anticardiolipin antibodies (IgG and IgM), lupus anticoagulant by activated partial thromboplastin time (APTT) and dilute Russell's viper venom test (DRVVT) and fasting plasma homocysteine. Results were considered abnormal only if two consecutive tests were outside the reference range for the local population. Because these tests are subject to widespread inter-laboratory variation, specific cut-off points for abnormal dynamic thrombophilia have deliberately not been given: local reference ranges were derived from the local normal population and cannot and should not be extrapolated to other settings.
Genetic studies were performed to look for specific mutations to methylenetetrahydrofolate reductase (MTHFR C677T), G20210A prothrombin gene and factor V Leiden.
Coagulation investigations
Antithrombin III, protein C activity, APCR without predilution with factor V deficient plasma, lupus clotting screens [prothrombin time (PT), APTT, DRVVT] with phospholipid neutralisation procedure (PNP) were all measured on the multidiscrete analyser (MDA-180 Organon-Teknika) with a variety of reagents according to the manufacturer's instructions. Protein S assay was performed using Diagnostica Stago free and total Protein S kits (Asnieres, France) by an enzyme-linked immunosorbent assay (ELISA) method.
Homocysteine and anticardiolipin antibodies
Homocysteine was quantified by ELISA using Axis Homocysteine EIA® (Bio-Rad, Hemel Hempstead, UK). Anticardiolipin antibodies were measured using the Cambridge Life Science Melisa kit (Cambridge Life Science, Huntingdon, UK). Results were read on multiscan reader (Titertek Multiscan II MCC/340®; from Laboratories Ltd, Finland) at 450 nm and calculated using the manufacturer's software.
DNA extraction and genetic analysis
Genomic DNA was extracted from 400 µl of whole blood using the Puregene kit® (Gentra, MN, USA) (floegen) according to the manufacturer's instructions. DNA sequences of factor V gene, prothrombin gene and MTHFR gene were amplified by the polymerase chain reaction (PCR) using primers (Bertina et al., 1994; Frosst et al., 1995
; Poort et al., 1996
).
Definition of placental lesions
The histological examination of the placenta was performed as a routine clinical practice in pregnancies with severe adverse outcome.
Fetal stem vessel thrombosis was diagnosed when a major placental vessel was completely or partially occluded by a thrombus. The vessel could be located either in the chorionic plate or in one of the stem vessel branches (arterial) or tributaries (vein).
Fetal thrombotic vasculopathy was identified by foci of more than five terminal villi showing lack of villous capillaries and with a hyalinized fibrotic stroma. The adjacent villi were normally vascularized and the distribution of avascular villi conformed to a single villous tree (Redline and Pappini, 1995).
Placental (villous) infarctions were diagnosed in the presence of an area of ischaemic necrosis of the placental villi with collapse of the intervillous space and tight clustering of the villous tree.
Perivillous fibrin deposition was diagnosed in the presence of increased and excessive accumulations of fibrin-like material in the intervillous space surrounding multiple villi that remained separated. Trophoblastic cells may have migrated into the fibrin deposits and the villous capillaries may be normal.
Intervillous thrombosis was diagnosed when a villous-free nodular thrombus was found in the intervillous space.
Placental floor infarction was diagnosed when the maternal surface of the placenta was extensively thickened and stiffened due to accumulation of excessive fibrinoid material which expands the basal plate, envelops anchoring villi and surrounds large numbers of basal and periseptal villi (Kraus, 1996).
Odds ratio (OR) and 95% confidence intervals (95% CI) were calculated for comparison of frequencies. Chi-square (2) test, Fisher's exact test, Student's t-test were used where appropriate and P < 0.05 was taken as significant. Statistical analyses were performed using the statistical package of Arcus Quickstat 1.0 (Iain Buchan, 1997).
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Results |
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Discussion |
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Placental infarction was the most common abnormal histological finding in our cohort and this finding is consistent with previous reports (Laurini et al., 1994). Small placental infarctions are common in uncomplicated pregnancies at term and of no importance, but extensive infarction, i.e. necrosis of >10% of the placenta, is associated with fetal hypoxia, growth retardation and fetal death, particularly in the second and early third trimester (Fox, 1997
; Rodriguez et al., 1997
).
Although it is claimed that fetal thrombotic vasculopathy and fetal stem vessel thrombosis are common findings in women with adverse pregnancy outcome (Arias et al., 1998), we were unable to confirm this observation.
The incidence of maternal floor infarction in unselected pregnancies is <1% (Naeye, 1985; Andres et al., 1990
). The incidence in our cohort was much higher (15%) confirming the previously observed association between maternal floor infarction of the placenta and adverse pregnancy outcome (Andres et al., 1990
). As expected, perivillous fibrin deposition and intervillous thrombosis were quite common, but their clinical significance remains questionable (Fox, 1998
).
Low activated protein C resistance (APCR) was the most common abnormal finding in this cohort. It is important to stress that only two women in our cohort had factor V Leiden mutation. Despite that, placental infarction was demonstrated in 50% of cases of low APCR ratio. The significance of this finding remains uncertain.
Placental changes in women with abnormal anticardiolipin antibodies and lupus anticoagulants are consistent with previous reports (Out et al., 1991; Magid et al., 1998
). Non-specific placental changes in women with hyperhomocysteinaemia were recently reported (Khong and Hague, 1999
) and were confirmed in our study. It is noteworthy that four women with hyperhomocysteinaemia had normal placentae. This suggests that placental changes may not be the only factor responsible for adverse pregnancy outcome in these women.
In summary, we found a poor correlation between thrombophilia status and pathological changes of the placenta in women with severe pregnancy complications. If there is a causal link between currently known thrombophilias, placental pathological changes and severe pregnancy complications, more sophisticated methods are required to prove it.
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Acknowledgments |
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Notes |
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References |
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Submitted on February 11, 2000; accepted on May 11, 2000.