93
Early Human Development, 23 (1990) 93-99 Elsevier Scientific Publishers Ireland Ltd. EHD 01075
Placental leukotriene B, release in early pregnancy and in term and preterm labour
A. Lbpez Bernal, D.J.
Hansell, T.Y. Khong, A.C. Turnbull
J.W. Keeling and
University of Oxford, Nuffield Department of Obstetrics and Gynaecology; John Radcliffe Hospital, Maternity Department, Headington. Oxford, OX3 9DU (U.K.) (Received 25 January 1990; revision received 10 April 1990; accepted 30 April 1990)
Summary
The production of leukotriene B, (LTB,) by the human placenta in vitro was measured by radioimmunoassay. In early pregnancy (7-12 weeks gestation) LTB, production rate (pmol/mg dry weight/2 h) was 3.7 (2.6-4.7) (median and range; n = 9) and at term it was 0.7 (0.4-2.3; n = 10) in placental tissue obtained at elective caesarean section, and 2.7 (1.3-3.6; n = 10) in samples following labour of spontaneous onset and vaginal delivery. In spontaneous preterm labour (26-36 weeks gestation) with normal placental histology LTB, production was 0.7 (0.31.7;n = 14), but it was significantly higher in preterm placentas with inflammatory infiltration: 3.1 (0.8-4.8; n = 6). These data show that the production of LTB, by human placenta is high in early pregnancy, but remains low during the third trimester, with a significant increase in spontaneous labour at term. LTB, output is low in uncomplicated preterm labour but markedly increased in chorioamnionitis-associated preterm labour. parturition; prematurity; infection; chorioamnionitis;
eicosanoids.
Introduction
Preterm labour remains a major cause of perinatal mortality and morbidity but its aetiology remains obscure. Leukotriene B, (LTB,) is a potent chemotactic and cell-activating agent involved in inflammatory and immune reactions; it is secreted following Slipoxygenase activation in polymorphonuclear leukocytes and macro0378-3782/90/$03.50 0 1990 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
94
phages [l], but may also be present in other cell and tissue extracts, including the human placenta, where its function is not well understood [lo]. LTB, levels increase in cord blood and in amniotic fluid following spontaneous labour at term [ 11,131, and in preterm labour associated with chorioamnionitis there is very high LTB, release by amniotic membranes with inflammatory infiltration [7]. This raises the possibility that LTB, measurements may be used as a marker for the presence of chorioamnionitis, and indeed raised amniotic fluid LTB, levels have been found in preterm labour associated with infection [ 141. The purpose of this paper was to compare the production of LTB, by human placenta obtained after uncomplicated and chorioamnionitis-associated preterm labour, and to study the influence of gestational age and the spontaneous onset of labour at term. Patients and Methods Placentas were obtained from 20 women who delivered following spontaneous preterm labour between 24 and 36 weeks gestation; from 10 women delivered before labour at term by elective caesarean section because of previous section, malposition or malpresentation; and from 10 women following spontaneous labour and vaginal delivery at term. All women had singleton pregnancies and further clinical data are given in Table I. Placental tissue was also obtained from nine women who had a surgical termination of pregnancy by vacuum aspiration at 7-12 weeks gestation. The study had the approval of the Central Oxfordshire Research Ethics Committee. The tissues were collected and rinsed in heparinised phosphate-buffered saline to remove blood, and were examined histologically in all cases. Blocks of placenta including the fetal and maternal surfaces, and samples of fetal membranes from the placental and peripheral part of the amniotic sac were cut, and a length of umbilical cord from the placental end was taken. The tissues were examined for polymorphonuclear leukocyte infiltration, other cellular infiltrates and evidence of inflammation. Histological examination was performed without knowledge of the leukotriene estimations. Fresh placental slices (about 100 mg wet weight), prepared from the central part of the organ after removing the decidual plate, were incubated in triplicate in a modified Krebs solution [7] at 37OC for 2 h. The viability of the tissue was checked by measuring glucose to lactate conversion. Preliminary experiments showed that under these conditions LTB, output was constant over a period of at least 3 h. After incubation, the medium was extracted with C,, minicolumns (JT Baker Chemical Co, Phillipsburgh, NJ) conditioned with methanol and water and eluted with water, hexane and methanol. The recovery of radiolabelled LTB, by this method was 8297%. The methanol extracts were dried down and redissolved in 0.04 M phosphate buffered saline (pH 7.4) containing 0.1 Vobovine gammaglobulin. Radioimmunoassay was performed essentially as described elsewhere [3] with anti-LTB, serum kindly donated by Dr R. Forder (ICI Bioscience, Macclesfield, U.K.). The specificity of this antiserum was: LTB, 100% cross reaction; all trans-LTB, isomer I 2.2%, isomer II 0.8%; 20-OH LTB, 0.3%; HHT 0.2%; 12s HETE, 12R/S HETE, 20-COOH
20(16-31)* 0(0-l) 10 (7-12) -
*Significant differences between groups (P< 0.01). j’P< 0.05 compared to uncomplicated preterm labour.
Maternal age (years) Parity Weeks gestation Apgar (5 min) Newborn weight (kg) Length of labour (h)
Early pregnancy (Ii = 9)
Pre-term labour
26 (17-39) 0 (O-2) 34 (31-36) 10 (S-10) 2.5 (1.5-2.7) 6.2 (2.5-10.1)
Uncomplicated (II = 14)
Clinical details of the patients. Results are medians and ranges.
TABLE I
29 (18-33) 1 (O-4) 34 (24-36) 9 (4-10) I .9 (0.6-2.7)t 4.7 (4.2-8.5)
Inflammatory (n = 6)
27 (16-32) 2 (o-3)* 39 (38-41) 10 (8-10) 3.3 (2.2-4.2) -
Elective LSCS (n = 10)
28 (20-40) 1 (O-3) 40 (38-41) 10 (9-10) 3.4 (2.7-4.5) 5.4(1.7-13.6)
Labour at term (n = 10)
96
LTB,, 6-oxo-PGF,., PGE,, TXB,, LTD, and arachidonic acid < 0.1% (data from Dr Forder). The recovery by radioimmunoassay of LTB, added to the incubation medium (5-20 pmol/ml) was 79-l 18%. The sensitivity of the standard curve was 15 fmol/tube. Intra- and interassay coefficients of variation were 9.3% and 13.2% (n = 5 for each). Statistical differences were assessed by the Kruskal-Wallis and Mann Whitney tests. Results Of the 20 patients delivered after spontaneous preterm labour, placental histology was normal in 14 patients and six had one or more of the following inflammatory changes: chorioamnionitis, acute or chronic deciduitis, intervillositis, funisitis, focal villitis. One of the patients had Listeria monocytogenes infection with aggregation of polymorphs within the intervillous space and formation of microabscesses throughout the depth of the placenta. For convenience this group of six patients will be referred to as the “chorioamnionitis” group. All term and early pregnancy specimens showed normal histology for their gestational age with no evidence of inflammation. The highest LTB, production rate (pmol/mg dry weight/2 h) was found in early pregnancy: 3.7 (2.6-4.3) (median and range) and the lowest at elective caesarean section at term, 0.7 (0.4-2.3), and after uncomplicated preterm labour, 0.7 (0.31.7) (P < 0.001). Individual values are plotted in Fig. 1. It can also be seen that the A
. 0. . t .
A
: .
.
A
t A
. 0
Early Pregnancy
;
A -
. PI A
-
Preterm ??Uncon ..
abour licated
. .
t
. .
.
0i
Elective LSCS
.
Labour at Term
Fig. 1. Placental leukotriene B, production rate in early pregnancy and in term and preterm labour with and without chorioamnionitis. The histograms represent median values.
91
spontaneous onset of labour at term was associated with a significant increase in LTB, output (P < 0.02 compared to the elective section group) and that the values in chorioamnionitis-associated preterm labour were considerably higher than in uncomplicated preterm labour (P < 0.001). Discussion This paper confirms that the human placenta is a source of immunoreactive LTB,. Ogburn et al. [lo] estimated LTB, by radioimmunoassay in extracts of term placenta and found levels of the order of l-2 pmol/mg dry weight. Experiments with radiolabelled arachidonic acid substrate [ 151 showed that first trimester placental cells produced compounds that co-eluted on HPLC systems with LTB, and mono-hydroxyeicosatetraenoic acids (HETEs), thus confirming the presence of 5lipoxygenase activity in the placenta. The conversion rates were higher in early pregnancy than in third trimester placental cells [15] and we have confirmed this pattern by radioimmunoassay. The relatively high LTB, output by early placental tissue could be related to the immune and vascular changes necessary for implantation and trophoblast invasion, because LTB, increases vascular permeability and enhances suppressor T cell formation [9,12]. Early placental villi have a larger mesenchymal core and more prominent cytotrophoblast than term villi [4] and differences in cell composition may be reflected in LTB, output. Moreover, first trimester samples often included decidualised endometrial cells together with placental villi. About 75% of early decidual cells are of bone marrow origin, mostly large granular lymphocytes [17], and it is possible that these cells contributed to “placental” LTB, output. It is also possible that the conditions of collection of early pregnancy tissue (prostaglandin to ripen the cervix and vacuum aspiration) may have increased abnormally the ability of the tissue to generate leukotrienes in vitro. Our data demonstrate that the spontaneous onset of labour at term is associated with increased placental LTB, output and this suggests that the placenta is the source of the raised LTB, levels found in cord blood [ 1l] and in amniotic fluid [13] of women in labour. The mechanism responsible for this increase is not known, but it is likely to be the result of 5-lipoxygenase and/or LTA and LTB synthase activation because the level of non-esterified arachidonic acid substrate in term placenta is relatively high [lo]. Similar increases in the production of HETEs by placenta and fetal membranes have been found following spontaneous labour at term [2,5,15]. It is of interest that high placental LTB, levels are present after spontaneous labour at term, but not after spontaneous preterm labour. Low amniotic fluid concentrations of LTB, and other lipoxygenase products such as 12- and 15-HETE have been found in women with uncomplicated preterm labour [ 141. Since prostaglandin metabolites in maternal plasma and in amniotic fluid are also low in preterm labour compared with labour at term [6,16], the evidence suggests that arachidonic acid metabolism in intrauterine tissues via cycle- and lipoxygenase pathways is different in term and preterm labour and indicates some “maturational” change towards term. The high LTB, release in placentas from chorioamnionitis-associated preterm
98
labour is in agreement with previous findings in amniotic tissue with inflammatory infiltration [7]. In this study, the highest LTB, output was found in tissue from a patient with Listeria monocytogenes infection with placental microabscesses, confirming that activated polymorphs are a major source of this compound. It is unlikely that LTB, release has any direct influence on uterine contractility in infection-associated preterm labour because LTB, has no oxytocic effect on human myometrium in vitro [8]. However, its potent chemotactic effect would amplify the inflammatory process and stimulate the release of prostaglandins, active peptides, histamine and platelet activating factor which could provoke uterine contractions. From a clinical point of view the possible use of LTB, estimations for the diagnosis of chorioamnionitis would be of interest. However this seems impractical at the present time, because assuming that all the excess LTB, produced by the placenta and the amnion in chorioamnionitis could reach the maternal circulation without being metabolised, its concentration would be below 1 pmol/ml of plasma and this could only be measured by very sensitive and laborious assays following HPLC purification. Significant differences in amniotic fluid LTB, concentrations have been described between women with uncomplicated- and infection associated-preterm labour [14], but there was considerable overlap in the values of the two groups and the amniotic fluid samples have to be obtained by transabdominal amniocentesis. It is likely that the measurement of 20-hydroxy, 20-carboxy and other leukotriene metabolites that may accumulate in maternal urine offers a more practical approach. Further studies are required to establish the cell types involved in the synthesis of LTB, and its metabolism by normal and infiltrated intrauterine tissues to clarify the possible role of this eicosanoid in pregnancy and labour. Acknowledgements Supported by Action Research for the Crippled Child. We are grateful to the Delivery Suite staff at the John Radcliffe Hospital for their help in the collection of samples and to B. Godman, Upjohn Ltd, for a gift of LTB, standard. References 1 2
3
4
5
Austen, K.F. and Soberman, R.J. (1988): Perspectives on additional areas for research in leukotrienes. Ann. N.Y. Acad. Sci., 524, xi-xxv. Bennett, P.R., Rose, M.P., Myatt, L. and Elder, M.G. (1987): Pre-term labor: Stimulation of arachidonic acid metabolism in human amnion cells by bacterial products. Am. J. Obstet. Gynecol., 156,649-655. Carey, F. and Forder, R.A. (1986): Radioimmunoassay of LTB, and 6-trans LTB,: analytical and pharmacological characterisation of immunoreactive LTB, in ionophore stimulated human blood. Prostag. Leukot. Med., 22,57-70. Dearden, L. and Ockleford, CD. (1983): Structure of human trophoblast: correlation with function. In: Biology of Trophoblast, pp. 70-100. Editors: Y.W. Loke and A. Whyte. Elsevier Science Publishers BV, Amsterdam. Kinoshita, K. and Green, K. (1980): Bioconversion of arachidonic acid and related compounds in human amnion. Biochem. Med., 23, 185-197.
99
6
I
8 9 10
11 12 13
14
15 16
17
Kragt, H., Noort, W.A.. de Zwart F.A. and Keirse, M.J.N.C. (1988): Can preterm birth be predicted by measuring PGF metabolites and bketo-PGF,* in preterm labour?. 1st European Congress on Prostaglandins in Reproduction (ECPR), Abstract 19, p. 30. Editors: P. Husslein and Ch. Egarter. Facultas Universitatsverlag, Vienna. L6pez Banal. A., Hansell, D.J., Canete Soler. R., Keeling, J.W. and Turnbull, A.C. (1987): Prostaglandins. chorioamnionitis and preterm labour. Br. J. Obstet. Gynaecol., 94. 1156-l 158. Lopez Bernal. A., Canete Soler, R. and Turnbull. A.C. (1989): Are leukotrienes involved in human uterine contractility? Br. J. Obstet. Gynaecol., 96. 11-16. Malathy, P.V., Cheng. H.C. and Dey, SK. (1986): Production of leukotrienes and prostaglandins in the rat uterus during peri-implantation period. Prostaglandins, 32,605-614. Ogbum, P.L., Roberts, J.D., Dassopoulos, T. and Walker, J.L. (1988): Leukotriene B, 6-ketoprostaglandin F,_, thromboxane B, and free arachidonic acid levels in human placental tissue. Ann. N.Y. Acad. Sci., X24.434-435. Pa&to, N., Piccione, E., Ticconi. C., Pontieri, G., Lenti, L. and Zicari. A. (1989): Leukotrienes in human umbilical plasma at birth. Br. J. Obstet. Gynaecol., %, 88-91. Rola-Pleszcxynski, M. (1985): Differential effects of leukotriene B, on T4+ and T8’ lymphocyte phenotype and immunoregulatory functions. J. lmmunol., 135,1357-1360. Romero, R., Emamian, M., Wan, M., Grzyboski, C., Hobbins, J.C. and Mitchell, M.D. (1987a): Increased concentrations of arachidonic acid lipoxygenase metabolites in amniotic fluid during parturition. Obstet. Gynecol., 70,849-851. Romero, R., Quintero, R., Emamian, M., Wan, M., Grzyboski, C.. Hobbins, J.C. and Mitchell, M.D. (1987b): Arachidonate lipoxygenase metabolites in amniotic fluid of women with intraamniotic infection and preterm labour, Am. J. Obstet. Gynecol., 157, 1454-1460. Rose, M.P. (1985): Arachidonic Acid Metabolism by Human Fetal Membranes in Tissue Culture, 132 pp. Ph.D. Thesis. University of London. Sellers, S.M.. Mitchell, M.D., Bibby, J.G., Anderson, A.B.M. and Turnbull, C.A. (1981): A comparison of plasma prostaglandin levels in term and preterm labour. Br. J. Obstet. Gynaecol., 88, 362-366. Starkey, P.M., Sargent, I.L. and Redman, C.W.G. (1988): Cell populations in human early pregnancy decidua: characterisation and isolation of large granular lymphocytes by flow cytometry. Immunology, 65, 129-134.
Early Human Development, 23 (1990) 93-99 Elsevier Scientific Publishers Ireland Ltd. EHD 01075
Placental leukotriene B, release in early pregnancy and in term and preterm labour
A. Lbpez Bernal, D.J.
Hansell, T.Y. Khong, A.C. Turnbull
J.W. Keeling and
University of Oxford, Nuffield Department of Obstetrics and Gynaecology; John Radcliffe Hospital, Maternity Department, Headington. Oxford, OX3 9DU (U.K.) (Received 25 January 1990; revision received 10 April 1990; accepted 30 April 1990)
Summary
The production of leukotriene B, (LTB,) by the human placenta in vitro was measured by radioimmunoassay. In early pregnancy (7-12 weeks gestation) LTB, production rate (pmol/mg dry weight/2 h) was 3.7 (2.6-4.7) (median and range; n = 9) and at term it was 0.7 (0.4-2.3; n = 10) in placental tissue obtained at elective caesarean section, and 2.7 (1.3-3.6; n = 10) in samples following labour of spontaneous onset and vaginal delivery. In spontaneous preterm labour (26-36 weeks gestation) with normal placental histology LTB, production was 0.7 (0.31.7;n = 14), but it was significantly higher in preterm placentas with inflammatory infiltration: 3.1 (0.8-4.8; n = 6). These data show that the production of LTB, by human placenta is high in early pregnancy, but remains low during the third trimester, with a significant increase in spontaneous labour at term. LTB, output is low in uncomplicated preterm labour but markedly increased in chorioamnionitis-associated preterm labour. parturition; prematurity; infection; chorioamnionitis;
eicosanoids.
Introduction
Preterm labour remains a major cause of perinatal mortality and morbidity but its aetiology remains obscure. Leukotriene B, (LTB,) is a potent chemotactic and cell-activating agent involved in inflammatory and immune reactions; it is secreted following Slipoxygenase activation in polymorphonuclear leukocytes and macro0378-3782/90/$03.50 0 1990 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
94
phages [l], but may also be present in other cell and tissue extracts, including the human placenta, where its function is not well understood [lo]. LTB, levels increase in cord blood and in amniotic fluid following spontaneous labour at term [ 11,131, and in preterm labour associated with chorioamnionitis there is very high LTB, release by amniotic membranes with inflammatory infiltration [7]. This raises the possibility that LTB, measurements may be used as a marker for the presence of chorioamnionitis, and indeed raised amniotic fluid LTB, levels have been found in preterm labour associated with infection [ 141. The purpose of this paper was to compare the production of LTB, by human placenta obtained after uncomplicated and chorioamnionitis-associated preterm labour, and to study the influence of gestational age and the spontaneous onset of labour at term. Patients and Methods Placentas were obtained from 20 women who delivered following spontaneous preterm labour between 24 and 36 weeks gestation; from 10 women delivered before labour at term by elective caesarean section because of previous section, malposition or malpresentation; and from 10 women following spontaneous labour and vaginal delivery at term. All women had singleton pregnancies and further clinical data are given in Table I. Placental tissue was also obtained from nine women who had a surgical termination of pregnancy by vacuum aspiration at 7-12 weeks gestation. The study had the approval of the Central Oxfordshire Research Ethics Committee. The tissues were collected and rinsed in heparinised phosphate-buffered saline to remove blood, and were examined histologically in all cases. Blocks of placenta including the fetal and maternal surfaces, and samples of fetal membranes from the placental and peripheral part of the amniotic sac were cut, and a length of umbilical cord from the placental end was taken. The tissues were examined for polymorphonuclear leukocyte infiltration, other cellular infiltrates and evidence of inflammation. Histological examination was performed without knowledge of the leukotriene estimations. Fresh placental slices (about 100 mg wet weight), prepared from the central part of the organ after removing the decidual plate, were incubated in triplicate in a modified Krebs solution [7] at 37OC for 2 h. The viability of the tissue was checked by measuring glucose to lactate conversion. Preliminary experiments showed that under these conditions LTB, output was constant over a period of at least 3 h. After incubation, the medium was extracted with C,, minicolumns (JT Baker Chemical Co, Phillipsburgh, NJ) conditioned with methanol and water and eluted with water, hexane and methanol. The recovery of radiolabelled LTB, by this method was 8297%. The methanol extracts were dried down and redissolved in 0.04 M phosphate buffered saline (pH 7.4) containing 0.1 Vobovine gammaglobulin. Radioimmunoassay was performed essentially as described elsewhere [3] with anti-LTB, serum kindly donated by Dr R. Forder (ICI Bioscience, Macclesfield, U.K.). The specificity of this antiserum was: LTB, 100% cross reaction; all trans-LTB, isomer I 2.2%, isomer II 0.8%; 20-OH LTB, 0.3%; HHT 0.2%; 12s HETE, 12R/S HETE, 20-COOH
20(16-31)* 0(0-l) 10 (7-12) -
*Significant differences between groups (P< 0.01). j’P< 0.05 compared to uncomplicated preterm labour.
Maternal age (years) Parity Weeks gestation Apgar (5 min) Newborn weight (kg) Length of labour (h)
Early pregnancy (Ii = 9)
Pre-term labour
26 (17-39) 0 (O-2) 34 (31-36) 10 (S-10) 2.5 (1.5-2.7) 6.2 (2.5-10.1)
Uncomplicated (II = 14)
Clinical details of the patients. Results are medians and ranges.
TABLE I
29 (18-33) 1 (O-4) 34 (24-36) 9 (4-10) I .9 (0.6-2.7)t 4.7 (4.2-8.5)
Inflammatory (n = 6)
27 (16-32) 2 (o-3)* 39 (38-41) 10 (8-10) 3.3 (2.2-4.2) -
Elective LSCS (n = 10)
28 (20-40) 1 (O-3) 40 (38-41) 10 (9-10) 3.4 (2.7-4.5) 5.4(1.7-13.6)
Labour at term (n = 10)
96
LTB,, 6-oxo-PGF,., PGE,, TXB,, LTD, and arachidonic acid < 0.1% (data from Dr Forder). The recovery by radioimmunoassay of LTB, added to the incubation medium (5-20 pmol/ml) was 79-l 18%. The sensitivity of the standard curve was 15 fmol/tube. Intra- and interassay coefficients of variation were 9.3% and 13.2% (n = 5 for each). Statistical differences were assessed by the Kruskal-Wallis and Mann Whitney tests. Results Of the 20 patients delivered after spontaneous preterm labour, placental histology was normal in 14 patients and six had one or more of the following inflammatory changes: chorioamnionitis, acute or chronic deciduitis, intervillositis, funisitis, focal villitis. One of the patients had Listeria monocytogenes infection with aggregation of polymorphs within the intervillous space and formation of microabscesses throughout the depth of the placenta. For convenience this group of six patients will be referred to as the “chorioamnionitis” group. All term and early pregnancy specimens showed normal histology for their gestational age with no evidence of inflammation. The highest LTB, production rate (pmol/mg dry weight/2 h) was found in early pregnancy: 3.7 (2.6-4.3) (median and range) and the lowest at elective caesarean section at term, 0.7 (0.4-2.3), and after uncomplicated preterm labour, 0.7 (0.31.7) (P < 0.001). Individual values are plotted in Fig. 1. It can also be seen that the A
. 0. . t .
A
: .
.
A
t A
. 0
Early Pregnancy
;
A -
. PI A
-
Preterm ??Uncon ..
abour licated
. .
t
. .
.
0i
Elective LSCS
.
Labour at Term
Fig. 1. Placental leukotriene B, production rate in early pregnancy and in term and preterm labour with and without chorioamnionitis. The histograms represent median values.
91
spontaneous onset of labour at term was associated with a significant increase in LTB, output (P < 0.02 compared to the elective section group) and that the values in chorioamnionitis-associated preterm labour were considerably higher than in uncomplicated preterm labour (P < 0.001). Discussion This paper confirms that the human placenta is a source of immunoreactive LTB,. Ogburn et al. [lo] estimated LTB, by radioimmunoassay in extracts of term placenta and found levels of the order of l-2 pmol/mg dry weight. Experiments with radiolabelled arachidonic acid substrate [ 151 showed that first trimester placental cells produced compounds that co-eluted on HPLC systems with LTB, and mono-hydroxyeicosatetraenoic acids (HETEs), thus confirming the presence of 5lipoxygenase activity in the placenta. The conversion rates were higher in early pregnancy than in third trimester placental cells [15] and we have confirmed this pattern by radioimmunoassay. The relatively high LTB, output by early placental tissue could be related to the immune and vascular changes necessary for implantation and trophoblast invasion, because LTB, increases vascular permeability and enhances suppressor T cell formation [9,12]. Early placental villi have a larger mesenchymal core and more prominent cytotrophoblast than term villi [4] and differences in cell composition may be reflected in LTB, output. Moreover, first trimester samples often included decidualised endometrial cells together with placental villi. About 75% of early decidual cells are of bone marrow origin, mostly large granular lymphocytes [17], and it is possible that these cells contributed to “placental” LTB, output. It is also possible that the conditions of collection of early pregnancy tissue (prostaglandin to ripen the cervix and vacuum aspiration) may have increased abnormally the ability of the tissue to generate leukotrienes in vitro. Our data demonstrate that the spontaneous onset of labour at term is associated with increased placental LTB, output and this suggests that the placenta is the source of the raised LTB, levels found in cord blood [ 1l] and in amniotic fluid [13] of women in labour. The mechanism responsible for this increase is not known, but it is likely to be the result of 5-lipoxygenase and/or LTA and LTB synthase activation because the level of non-esterified arachidonic acid substrate in term placenta is relatively high [lo]. Similar increases in the production of HETEs by placenta and fetal membranes have been found following spontaneous labour at term [2,5,15]. It is of interest that high placental LTB, levels are present after spontaneous labour at term, but not after spontaneous preterm labour. Low amniotic fluid concentrations of LTB, and other lipoxygenase products such as 12- and 15-HETE have been found in women with uncomplicated preterm labour [ 141. Since prostaglandin metabolites in maternal plasma and in amniotic fluid are also low in preterm labour compared with labour at term [6,16], the evidence suggests that arachidonic acid metabolism in intrauterine tissues via cycle- and lipoxygenase pathways is different in term and preterm labour and indicates some “maturational” change towards term. The high LTB, release in placentas from chorioamnionitis-associated preterm
98
labour is in agreement with previous findings in amniotic tissue with inflammatory infiltration [7]. In this study, the highest LTB, output was found in tissue from a patient with Listeria monocytogenes infection with placental microabscesses, confirming that activated polymorphs are a major source of this compound. It is unlikely that LTB, release has any direct influence on uterine contractility in infection-associated preterm labour because LTB, has no oxytocic effect on human myometrium in vitro [8]. However, its potent chemotactic effect would amplify the inflammatory process and stimulate the release of prostaglandins, active peptides, histamine and platelet activating factor which could provoke uterine contractions. From a clinical point of view the possible use of LTB, estimations for the diagnosis of chorioamnionitis would be of interest. However this seems impractical at the present time, because assuming that all the excess LTB, produced by the placenta and the amnion in chorioamnionitis could reach the maternal circulation without being metabolised, its concentration would be below 1 pmol/ml of plasma and this could only be measured by very sensitive and laborious assays following HPLC purification. Significant differences in amniotic fluid LTB, concentrations have been described between women with uncomplicated- and infection associated-preterm labour [14], but there was considerable overlap in the values of the two groups and the amniotic fluid samples have to be obtained by transabdominal amniocentesis. It is likely that the measurement of 20-hydroxy, 20-carboxy and other leukotriene metabolites that may accumulate in maternal urine offers a more practical approach. Further studies are required to establish the cell types involved in the synthesis of LTB, and its metabolism by normal and infiltrated intrauterine tissues to clarify the possible role of this eicosanoid in pregnancy and labour. Acknowledgements Supported by Action Research for the Crippled Child. We are grateful to the Delivery Suite staff at the John Radcliffe Hospital for their help in the collection of samples and to B. Godman, Upjohn Ltd, for a gift of LTB, standard. References 1 2
3
4
5
Austen, K.F. and Soberman, R.J. (1988): Perspectives on additional areas for research in leukotrienes. Ann. N.Y. Acad. Sci., 524, xi-xxv. Bennett, P.R., Rose, M.P., Myatt, L. and Elder, M.G. (1987): Pre-term labor: Stimulation of arachidonic acid metabolism in human amnion cells by bacterial products. Am. J. Obstet. Gynecol., 156,649-655. Carey, F. and Forder, R.A. (1986): Radioimmunoassay of LTB, and 6-trans LTB,: analytical and pharmacological characterisation of immunoreactive LTB, in ionophore stimulated human blood. Prostag. Leukot. Med., 22,57-70. Dearden, L. and Ockleford, CD. (1983): Structure of human trophoblast: correlation with function. In: Biology of Trophoblast, pp. 70-100. Editors: Y.W. Loke and A. Whyte. Elsevier Science Publishers BV, Amsterdam. Kinoshita, K. and Green, K. (1980): Bioconversion of arachidonic acid and related compounds in human amnion. Biochem. Med., 23, 185-197.
99
6
I
8 9 10
11 12 13
14
15 16
17
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