Maternal plasma fibronectin: a predictor of preterm delivery

ELSEVIER

European Journal of Obstetrics & Gynecology and Reproductive Biology 72 (1997) 121 -126

GYNECi)U

Maternal plasma fibronectin: a predictor of preterm delivery Marek Zygmunt ~, Uwe Lang ~, Norbert Katz b Wolfgang Kiinzel ~'* Department o/ Obstetrics/Gynaecology, University Of Giessen, Klinikstrafle 32, 35385 Giessen, Germany b Department ()f Clinical Chemistry, UniversiO' ~/' Gies~'en, Kl#~ikstrafie 32, 35385 Giessen, Germany Received 22 May 1996: received in revised form 20 June 1996: accepted 12 November 1996

Abstract

Objective: Current opinion holds that there are several distinct groups among patients with preterm labour: one of them is characterized by bacterial infection, another one by the presence of placental vascular abnormalities with endothelial damage. The aim of this study was to investigate plasma fibronectin, a suspected biochemical marker of endothelial damage, as an indicator for pregnancies with a high risk of preterm delivery. Methods: Plasma fibronectin levels were measured in patients with preterm labour (n = 80) and in healthy pregnant women with uncomplicated (control) pregnancies (n = 64) between the 22nd and 36th week of gestation. Furthermore, the plasma concentrations of fibronectin in 15 newborns at term and ten babies born preterm were measured to study the relationship between preterm delivery and plasma fibronectin concentration in newborns. Fibronectin was measured by nephelometry. Results: The mean concentration of fibronectin in patients with preterm labour was 0.44 g/1 (S.D., 0.15) vs. 0.25 g/l (S.D., 0.12) in uncomplicated control pregnancies matched for gestational age, In control patients who actually delivered at term, fibronectin values were found to be lower than in control patients who underwent preterm delivery (0.25 g/l: S.D., 0.05: vs. 0.46 g/l; S.D., 0.15; P < 0.05). Particularly high values were detected in patients with preterm labour delivering before 32 weeks of gestation (0.60 g/l; S.D., 0.16). There was no significant difference between fibronectin concentrations in the umbilical arterial and venous blood of premature infants and mature infants. Leucocyte concentration, bacteriological smear and cervical dilatation did not correlate with fibronectin concentrations in patients with preterm delivery or controls. Conclusion: We conclude that the higher plasma concentrations of fibronectin in women with preterm labour may be a biochemical marker and a predictor of preterm delivery. © 1997 Elsevier Science Ireland Ltd. Keywords: Plasma fibronectin; Preterm labour; Endothelial damage: Predictor of preterm delivery

1. Introduction

Preterm labour and premature delivery are still major problems o f preventive medical care today. The incidence o f preterm delivery is about 7 - 1 2 % o f all deliveries [1]. This fraction o f the infant population needs sophisticated medical care and causes high economical costs. M a n y theories [2-4] to explain the pathogenesis o f preterm labour and delivery have been created. Current data show that there seem to be several distinct

* Corresponding author: Tel.: +49 641 9945100: fax: +49 641 9945109.

groups a m o n g patients with preterm labor: one o f them is characterized by bacterial infection [5], a second one by presence o f placental vascular abnormalities, i.e. absence o f adaptative changes in spiral arterioles accompanied by multiple t h r o m b i and endothelial damage [6-8]. Those placental abnormalities have been described previously in gestosis, I U G R and pre-eclampsia. M a n y researchers have attempted to identify w o m e n more likely to be delivered before term [9-14]. The purpose o f our study was to investigate the use o f suspected systemical markers o f placental pathology and vascular endothelial d a m a g e (e.g. fibronectin) [15] as an indicator for preterm delivery.

0301-2115/97/$17.00 ~! 1997 Elsevier Science Ireland Ltd. All rights reserved. PII S0301-211 5( 96 )02671- 1

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M. Zvgmunt el al. European Journal O/Obstetrics & Gynecology and Reprmh~ctive Biology 72 (1997) 121 126

Table 1 Characteristics of the study groups Preterm labour

Control group

(n =

64)

Significance

07- 80)

Mean (S.D.), range

Mean (S.D.), range

(P<0.5)

Age (years)

28 (5) 18 41

28 (4.5) 17 39

NS

Estimated fetal weight at the date of ultrasound (g)

1775 (620)

1675 (690)

NS

Birthweight (g)

2900 (790)

3525 (510)

S

Gestational age at inclusion into the study (days/completed weeks)

217 (21) /31

221 (25) /31

NS

Gestational age at delivery (days/completed weeks)

262 (19) /37

280 (12) /40

S

Age of patients, gestational age at the time of inclusion in the study and at delivery and fetal weight estimated by ultrasound at delivery are presented as mean + S.D. P<0.05 was considered significant.

The present paper deals with the following questions: 1. is there any relationship between plasma levels of fibronectin and preterm labour? 2. are plasma concentrations of fibronectin a predictor of preterm labour and delivery? 3. is there any relationship between plasma levels of fibronectin and common markers of inflammation (e.g. C-reactive protein, leucocytosis)? 4. are plasma concentrations of fibronectin associated with premature or mature cervical ripening?

2. Patients and methods

This prospective study was performed over l0 months at the Department of Obstetrics and Gynaecology, University of Giegen (Germany). Preliminary investigations were performed at the Department of Obstetrics and Gynaecology, University of Aachen (Germany). Eighty patients admitted to the hospital with signs of preterm labour between 22 and 36 weeks of gestation were included into the study. All patients signed declarations of informed consent before entry into the study. Inclusion criteria were: 22- 36 weeks of gestation premature uterine contractions with a frequency of > 5/30 min in CTG-registration premature uterine contractions with a frequency of > 15 per day felt by the patient signs of premature cervical ripening (cervical change) premature rupture of membranes Patients with intrauterine fetal growth retardation, diabetes mellitus, fetal malformation, gestosis, liver disorders and coagulopathy (conditions with increased or decreased fibronectin levels) were excluded. The control

group consisted of 64 healthy pregnant women with uncomplicated pregnancy of 22 36 weeks of gestation who were outpatients of the Department of Obstetrics and Gynecology, University of Giegen. Women in the control group received no tocolytic agent such as flmimetics or indomethacin. Most, however, had oral medication with magnesium sulfate. In case of preterm labour steroid administration to accelerate fetal lung maturation took place after blood sampling for our study. To investigate whether uterine contractions exert a direct influence on plasma fibronectin concentrations an additional 20 healthy pregnant women without contractions were examined on their estimated day of delivery and for the second time in active labor with regular contractions and a cervical dilatation of 5 cm. Furthermore, the plasma concentrations of fibronectin in 15 newborns at term and ten babies born preterm were measured to study the relationship between preterm delivery and plasma fibronectin concentration (artery and vein blood) in newborns (Table 3). Table 2 Plasma fibronectin concentrations in non-pregnant women, pregnant patients with preterm labour and healthy pregnant women

1 2 3

Patient group

Fibronectin (g/l) mean (S.D.)

Non-pregnant ( n - 20) Healthy pregnant controls ( n 64) Pregnant with preterm labour (;7 - 80)

0.27 (0.05) 0.25 (0.12) 0.44 (0.15)

The differences between non-pregnant women (1) and patients with preterm labour (3) and between healthy pregnant women and patients with preterm delivery (3) were considered statistically significant (P<0.05).

M. Zvgmunt eta/.

European Journal of Ob.~tetrics & Qvneco/ogv amt Ru.'o&*ctive Biology 72 (1997) 121 126

Table 3 Plasma fibronectin concentrations arterial and venous blood) in children born at term and infants born preterm Patient group

Vessel

Fibronectin {g,'l) mean (S.D.)

Children born at term ( n - 15)

A. umbilicalis

0.19 ((I.05)

V. umbilicalis A. umbilicalis

0.20 (0.03) 0.16 (0.04)

V. nmbilicalis

0.19 (0.03)

Children born preterm ( n - 10)

There wets no significant differencebetween both groups.

We also measured plasma fibronectin concentration of 20 healthy non-pregnant women to compare them with values of pregnant women (Table 2). Blood samples were collected by venipuncture (5 ml) and placed in plastic tubes containing EDTA. After centrifugation (5000 rpm for 10 min) plasma was frozen in 40°C until assay. Quantitation of fibronectin was performed by nephelometry (Behring Marburg, Germany). Gestational ages were calculated from the last menstruation and confirmed by early ultrasound. When the dates were uncertain they were modified according to early ultrasonography. Cervical length was examined manually (Bishop score). A mature cervix was characterized by a Bishop score of greater than 4 points in nulliparous women and a Bishop score of greater than 5 points in multiparous women. Intrauterine infection and inflammatory response to infection was defined as a C-reactive protein concentration above 5 mg/l or leucocytosis above 10 000/#1 or a positive microbiological culture from the cervix. Fetal weight was estimated by ultrasonographic biometry.

3. Statistics

The relation between quantitative parameters was tested by multiple regression analysis. Significance of the differences between qualitative parameters was performed by ,g2 tests and contingency tables. To compare differences between groups, one and two-way analysis of variance was used. A P value of less than 0.05 was considered statistically significant. Sensitivity, specificity, positive and negative predictive values were also calculated. Data are expressed as mean (_+ S.D.).

4. Results

Characteristics of the study groups are summarized in Table 1. The study groups were similar with regard

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to age, gestational age at inclusion in the study as well as estimated fetal weight. Mean plasma concentration of fibronectin measured in pregnant women with preterm labour (n = 80) was 0.44 g/l (S.D., 0.15). Mean plasma concentrations of fibronectin in pregnant healthy control women (tl = 64) was signifcantly lower at 0.25 g/1 (S.D., 0.12) (Table 2). Gestational age was not related to fibronectin concentrations in both groups (Fig. 1). In control patients with subsequent preterm delivery (t7 = 6) plasma fibronectin concentrations were significantly higher (0.46 g/l: S.D., 0.15) than in control patients who delivered at term (0.25 g/'l; S.D., 0.05). Particularly high values were detected in four patients m the group of preterm labour who d'elivered before 32 weeks of gestation (0.60 g:l; S.D., 0.16) (Fig. 2). There was no correlation between plasma fibronectin concentrations and cervical dilatation (Bishop score). The mean concentrations of fibronectin in patients with a high Bishop score ( > 4 points ill nulliparous and > 5 points in multipara) were equivalent to values in patients with a low score (nulliparous women, 0.42 g,1, S.D. 0.15 vs. 0.45 g/l; S.D. 0.15: multiparous women, 0.39 g:l, S.D. 0.14; vs. 0.38 g/l, S D . 0.10: P > 0.05). Plasma fibronectin concentrations showed no increase with uterine contractions (0.32 g/l, S.D. 0.06; and 0.34 g/l, S.D. 0.05; P > 0.05). The mean concentrations of plasma fibronectin in patients with c-reactive protein above 5 mg/1 was equivalent to the plasma fibronectin concentration in the patients with c-reactive protein below 5 mg/1 (0.43 gl, S.D. 0.21 vs. 0.43 g/l, S.D. 0.15; P > 0 . 0 5 ) . No correlation between leucocyte count, bacteriological smear and the fibronectm concentration in palients with preterm delivery and controls could be established. There was no significant difference between the umbilical arterial and venous blood fibronectin concentrations from premature and mature infants (Table 3).

5. Fibronectin as predictor of preterm delivery

With or without preterm labour 90% of patients delivering alter 37 weeks of gestation had a fibronectin level of less than 0.45 g/l. Seventy-eight percent of all patients with a fibronectin level of 0.45 g/l or higher delivered before 37 weeks of gestation. Choosing a cut off level of 0.45 g/1 (mean of the control g r o u p + 2 S.D.) for a screening test gives a good specificity of 95% (per definitionem) and a poor sensitivity of about 50%. A positive predictive value of 50% (of preterm delivery if fibronectin > 0.45 g/l) and a negative predictive value of 90% was estimated in this study (Fig. 2).

124

M. Zygmunt et al./European Journal (~f Obstetrics & Gynecoh)gy and Reproductive Biology 72 (1997) 121 126 Fn[g/I] Preterm labour (n=80) ? + C o n t r o l (n=64)

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Fig. 1. Plasma concentrations of fibronectin [Fn] in patients with preterm labour (n = 80) and in controls (n - 64) sampled at a de[ned gestational age are shown. There was no significant correlation between the fibronectin concentration and gestational age in either group. The r value was 0.22 in the patients with preterm labour and 0.10 in controls.

Moreover, there was no significant difference between the umbilical blood fibronectin concentrations from premature and mature infants as has been shown by others [18]. We conclude that the higher fibronectin values in preterm labour are probably not an acute phase mediator or marker of localized infection (of the lower genital tact or chorionamionitis). Results of this study can be explained with in vitro experiments from Yoder et al. (1988) [19] and Jackson et al. (1993) [20] which show a lack of influence of prostaglandin inhibitors and corti-

6. Discussion

In the present study it was shown that plasma fibronectin values in women with preterm labour are significantly higher than in healthy pregnant women of the same gestational age. Plasma fibronectin concentrations in healthy pregnant controls delivering preterm are also significantly higher than in pregnant controls delivering at term. No correlation between gestational age and fibronectin concentration was found, which is in correspondence with previous investigations [16,17].

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+ Control

(n=64)

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Fig. 2. Fibronectin as a predictor of preterm delivery. Plasma fibronectin concentrations [Fn] as a random sample during gestation in relation to the week of delivery. Patients who delivered preterm had a higher average fibronectin concentration at the time of sampling than patients who delivered at term.

M. Zvgmunt et al. European Jourmd o/ Obstetrics & Gvnecolo~,Lv am/Repn)~h~ctive Biology 72 (1997) 121 126

son on t h e f i b r o n e c t i n b i o s y n t h e s i s in cell c u l t u r e . O n the o t h e r h a n d it was s h o w n by G u l l e r et al. (1995) [21] t h a t s t e r o i d a p p l i c a t i o n in a m n i o n cell c u l t u r e r e d u c e d the synthesis o f f i b r o n e c t i n . R o b e r t et a]. (1988) [22] a n d G o y e r t et al. (1990) [23] h a v e s h o w n t h a t p l a s m a levels o f f i b r o n e c t i n w e r e not s i g n i f c a n t l y d i f f e r e n t in patients with chorionamnionitis, patients with PROM and undelivered patients without antenatal complicat i o n s m a t c h e d for g e s t a t i o n a l age. A lack o f a r e l a t i o n ship b e t w e e n p l a s m a f i b r o n e c t i n v a l u e s a n d cervical r i p e n i n g suggests t h a t the h i g h f i b r o n e c t i n v a l u e s in p r e t e r m l a b o u r are n o t c a u s e d by release o f f i b r o n e c t i n f r o m the cervix uteri. S t u b b s et al. (1984) [15] p o s t u l a t e d t h a t the i n c r e a s e d p l a s m a f i b r o n e c t i n is the result o f release f r o m d a m a g e d e n d o t h e l i a l cells. T h e d a t a o f o u r c u r r e n t s t u d y c o u l d be e x p l a i n e d as e v i d e n c e o f e n d o t h e l i a l d a m a g e in p r e t e r m l a b o r . Salafia et al. (1991) [7] a n d A r i a s et al. (1993) [8] h a v e s h o w n e v i d e n c e o f m a t e r n a l p l a c e n t a l vasculopathy among patients with preterm labour. The c h a n g e s d e s c r i b e d by t h o s e i n v e s t i g a t o r s are similar to t h o s e f o u n d in I U G R , p r e - e c l a m p s i a o r h y p e r t e n s i o n d u r i n g p r e g n a n c y [24 28]. T h e y are c h a r a c t e r i z e d by lack o f a d a p t a t i v e c h a n g e s in the spiral arterioles, by p l a c e n t a l i n f a r c t i o n s a n d t h r o m b o s i s a c c o m p a n i e d by endothelial damage. T h e s e c h a n g e s also i n d i c a t e t h a t m e a s u r e m e n t o f f i b r o n e c t i n in m a t e r n a l p l a s m a c a n be useful for p r e d i c t i o n a n d p r o g n o s i s o f p r e t e r m d e l i v e r y if a cut o f f level o f 0.45 g/1 is c h o s e n a n d o t h e r p r e g n a n c y c o m p l i c a t i o n s i n f l u e n c i n g the f i b r o n e c t i n v a l u e s ( I U G R , d i a b e t e s , hyp e r t e n s i o n ) are e x c l u d e d . L o n g i t u d i n a l population b a s e d studies are r e q u i r e d to e x a m i n e at w h a t t i m e the f i b r o n e c t i n c o n c e n t r a t i o n in p a t i e n t s w i t h p r e t e r m l a b o u r starts to rise.

Acknowledgements W e t h a n k Mr. P a b s t ( D e p a r t m e n t o f Biostatistics, U n i v e r s i t y o f G i e s s e n , S c h o o l o f M e d i c i n e ) f o r the statistical analysis a n d m a n y useful s u g g e s t i o n s .

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