British Journal of Obstetrics and Gynaecology October 1976. Vol83. pp 115-779

PLASMA LEVELS OF PREGNANCY-SPECIFIC & -GLYCOPROTEIN IN NORMAL PREGNANCY BY

C . M. TOWLER, Research Fellow in Pathology

C . H. W. HORNE,Senior Lecturer in Pathology Department of Pathology, University Medical School, Foresterhill, Aberdeen V. JANDIAL, Senior Registrar in Obstetrics and Gynaecology DORISM. CAMPBELL, Lecturer in Obstetrics, Gynaecology and Physiology AND

I. MACGILLIVRAY, Regius Professor of Obstetrics and Gynaecology Departtnent of Obstetrics and Cynaecolog,y, Aberdeen Maternity Hospital Foresterhill, Aberdeen Summary Pregnancy-specific &-glycoprotein (PSPG) is a major product of the trophoblast which has only recently been identified. Like human placental lactogen, the concentration of PSPG in maternal blood rises throughout pregnancy until about 34 weeks, thereafter tending to remain constant until term, with only a small day-to-day variation in individuals. The circulating maternal levels of PSgG between 34 weeks and term are about 200 pg/ml, 20 to 30 times greater than the levels of placental lactogen, thus allowing detection and measurement of PSPG by relatively simple techniques. The levels of PSPG are above the normal range in the majority of twin pregnancies. It is suggested that PSPG measurements may be useful in assessing placental function and may help in the detection of multiple pregnancies at an early stage of gestation. ADEQUATE antenatal care depends on the early detection of any condition which may threaten the well-being of the fetus. In recent years, considerable attention has been paid to steroid and protein products of the trophoblast whose concentration in plasma or urine gives an indication of placental function and may thus be useful in identifying the ‘at risk’ fetus. A trophoblast product which is, as yet, unfamiliar to many clinicians is pregnancy specific PI-glycoprotein (PSBG), first described in 1971 by Bohn and referred to by him as SP1 (Bohn, 1971). This globulin which is not normally detectable in non-pregnant individuals is produced by the syncytiotrophoblast very soon

after implantation of the ovum (Horne el al, 1976) and is secreted into the maternal circulation where it has a half life of 30 to 40 hours (Bohn, 1974; Tatra et al, 1975). The relatively high plasma concentration of PSPG in the second half of pregnancy facilitates its measurement by simple laboratory techniques. At first sight, therefore, PSPG appears to merit consideration as an indicator of placental function since at least three of the essential criteria for such a test, as proposed by Chard (1974), are fulfilled. Before the prognostic value of PSPG measurements in assessing placental function can be evaluated, it is necessary to establish the range of values found in normal pregnancies and the 175

776

TOWLER, HORNE, JANDIAL, CAMPBELL AND MACGILLIVRAY

day-to-day variations in individuals. In this study, we have made such measurements and compared them with those of another trophoblast product, human placental lactogen (HPL) which arenow accepted as an indexof placentalfunction. Since it has been shown that HPL measurements may be a useful adjunct to ultrasonography in early diagnosis of multiple pregnancies (Grennert et al, 1976), we have studied plasma PSgG concentrations in both singleton and twin pregnancies. METHODS Plasma PSPG concentrations in normal pregnancy were studied between 24 and 41 weeks gestation in a group of 40 women (mean age 24; range 18 to 36) who were judged prospectively to be normal and who developed no antenatal complications. Thirty-three of the normal group here priniigravidae. Twenty-four subjects were studied serially, blood samples being obtained at weekly or fortnightly intervals. At least two blood samples were obtained from the remaining subjects. In all, about 250 venous blood samples were collected. Separated plasma was stored at -20 "C. Day-to-day variations in plasma PSpG and HPL concentrations were studied in six women from the normal group who gave a blood sample at the same time of day on six successive days. Plasma PSpG concentrations in twin pregnancies were studied in a group of nine subjects. Twins were suspected clinically and confirmed by ultrasonography. Plasma concentrations of PSPG and HPL were determined using the appropriate 'Partigen' immunodiffusion plates and standard solutions (Behringwerke AG, Germany). Plasma samples from twin pregnancies were diluted two-fold with 0 .9 per cent saline before assay. To minimise interplate variation, samples from any one patient were assayed on the same plate (Thompson et al, 1969). The coefficient of variation for 12 replicate assays on one plate was 2 .3 per cent for PSPG and 2.8 per cent for HPL.

RESULTS The plasma concentrations of PSSG and HPL (Mean &SD) from 24 weeks to term in

the normal pregnant subjects are shown in Table I and Figure 1. TABLE I Normal singleton ranges of PSpG and HPL ~

Week from last menstrual period 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41

Plasma PSpG concentrations (rgiml)

cv

Mean

SD

(per cent)

61.1 90.0 85.9 94.0 114.3 121.0 129.5 147.8 158.3 177.1 163.2 190.7 195.9 198.4 199.3 199.0 209.1 188.3

17.5 30.0 22.6 27.2 20.8 40.5 30.1 47.0 30.4 45.2 40.9 53.3 51.9 48.0 48.6 53.9 66.7 68.4

28.7 33.0 26.3 29.0 18.2 33.4 23.2 31.7 19.2 25.5 25.0 28.0 26.5 24.2 24.4 27.1 31.9 36.3

No. of subjects 7 7 10 6 9 8 11

12 12 13 15 11 20 18

21 16 15 8

mean 27.3 ~-

Week from last menstrual period 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41

-

Plasma HPL concentrations (IIg/ m 1) _____.- _____

cv

Mean

SD

(per cent)

2.70 3.27 3.23 4.10 4.64 5.33 5.16 6.23 6.16 7.52 7.26 7.96 7.60 7.86 7.65 7.06 7-20 7.61

1.21 1.14 1.08 0.41 1.94 0.99 1.14 0.71 1.42 1.35 1.60 1.76 1.66 1.69 1.70 I .71 1.74 2.53

45.0 34.9 33.4 10.0 41.8 18.7 22.1 11.4 23.1 18.0 22.0 22.2 21.8 21.5 22.2 24.2 24.1 33.3

mean 25.0

No. of subjects

5

7 10 6 8 8 11 10 15 10 12 14 20 18 22 19 14 8

PSgG IN NORMAL PREGNANCY 350

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38

40

Week of Gestation

FIG.1 The ranges of plasma PSPC and HPL concentrations found in 40 normal singleton pregnancizs. (SD standard deviation).

Since there was no obvious difference between the levels of these proteins in primigravid and parous women, all values were used in constructing these graphs. The shapes of the curves are similar, the mean concentrations of both proteins rising continuously until around 34 weeks, thereafter levelling out with a tendency for the concentrations (particularly of HPL) to fall during the last few weeks of pregnancy. In most cases, plots of serial PSgG and HPL values from individuals were of a similar shape to the mean curves for the total sample. Despite these similarities, there was no correlation between plasma PSgG and HPL concentrations (r = -0.18; n = 60).

The day-to-day variations of PSPG and HPL in six normal pregnant subjects over six consecutive days are shown in Figure 2. Whereas the mean coefficient of variation for the PSPG values was 5.7 per cent, that for HPL was 4.4 per cent. These coefficients of variation are only slightly higher than the experimental variation incurred in carrying out the estimations. As shown in Figure 3, levels of both PSgG and HPL were raised above the normal range in normal twin pregnancies, all values falling above the mean +ISD. In many cases, the values fell above the upper limit (mean +2SD) of the normal range.

778

TOWLER, HORNE, JANDIAL, CAMPBELL AND MACGILLIVRAY Day of sampling dear

S.D.

167.t 5.5

1 t

:.V. 1.3%

6.5 7.5 196

14.3

Day of sampling 2 3 4 5 , r , ,

7.3%

6.5 -

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C.V.

1.47

1.47

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5.88

0.33

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10.52

0.33

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5.98

0.31

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7.27

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6.89

0.25

3.7%

5.5

164

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w 170 4

z

147.

13.8

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9.9

6.7%

-

10.0

5.0

140

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4.1

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mean C.V.

9.8

L

4.2%

6.f

-

6.0

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-'--

mean C.V.

4.4%

FIG.2 Day-to-day variations of the plasma PSPG and HPL concentrations in six women between 32 and 40 weeks gestation.

DISCUSSION Chard (1974) has proposed that for any plasma constituent to be a useful monitor of placental function four criteria should be satisfied. PSgG would appear to satisfy these criteria since (1) it is not normally present in non-pregnant subjects, (2) it shows only a small day-to-day variation, (3) its half life is relatively short, being approximately 30 hours calculated from its rate of disappearance post partum, (4) the plasma concentrations are sufficiently high to be readily measured by any of the rapid established techniques such as Laurel1 eIectroimmunodiffusion and, of course, radioimmunoassay. We submit, therefore, that measurements of PSgG may prove to be useful in assessing placental function, particularly in view of the finding that plasma PSgG levels correlate with placental weight (Tatra et al, 1975) and appear to fall outside the normal range in some of the few abnormal pregnancies reported to date (Tatra et al, 1974). While our normal range for HPL is similar to

that reported by Singer et a1 (1970) and only slightly different to those of other groups (Genazzani et al, 1971 ; Radiochemical Centre, 1973), there are few studies with which we can compare our normal PSgG range. Tatra et al (1974), in a comparable study to our own, have shown a graph for the normal range for PSPG whose shape is similar to ours. In addition, the scatter of values in their range (mean coefficient of variation = 28.5 per cent) was similar to that in ours (mean CV = 27.3 per cent). However, the mean values of the plasma PSSG concentrations in the second half of pregnancy as reported by Tatra et a1 (1974) were more than 30 per cent lower than our mean values. Although the reason for this discrepancy is not clear, the use of different standard PSPG preparations may be an important factor. The range of plasma PSgG values observed in a previous study of 15 normal primigravidae (Towler et al, 1976) was similar to that reported here. A recent report (Grennert et al, 1976) has

PSgG IN NORMAL PREGNANCY 350-

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Weak of Gatmian

779

may be raised above the normal singleton range in conditions other than inultiple pregnancy. We propose that raised levels of both PSpG and HPL may give a more reliable indication of multiple pregnancy than a high plasma level of only one of the two proteins. Since both PSpG and HPL are produced by the trophoblast very early in pregnancy, with suitably sensitive assay techniques the diagnosis of twins may be possible in the first trimester of pregnancy. These observations, together with preliminary evidence that PSpG levels may be useful in detecting intrauterine growth retardation of the fetus amongst patients with severe pre-eclampsia (Jandial et al, 1976) suggest that PSpG merits serious consideration as a possible indicator of placental function.

ACKNOWLEDGEMENTS The authors thank the Medical Research Council for financial support and Behringwerke AG, MarburgILahn, West Germany who kindly provided the Partigen plates and protein standards. We are also indebted to Miss Lilian Hopkins for her excellent technical assistance.

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Week of Gestation

FIG.3 The plasma concentrations of PSPG and HPL in nine twin pregnancies. The shaded areas represent the limits (mean &2SD) of thenormal rangein singleton pregnancies.

suggested that measurements of HPL may be a useful alternative to ultrasonography in the early diagnosis of twins. It is possible that PSpG measurements may also be useful in this way since we have shown that PSpG levels are above the normal range in the majority of twin pregnancies. However, HPL or PSpG levels

REFERENCES Bohn, H. (1971): Archiv fur Gynakologie, 210,440. Bohn, H. (1974): Archivfiv Gynukologie, 216, 347. Chard, T. (1974): Clinics 6 2 Obstetrics and Gynaecology, 1, 85. Genazzani, A. R., Cocola, F., Casoli, N., Mello, G., Scarselli, G., Neri, P., and Fioretti, P. (1971): Journal OJ Obstetrics and Cynaecology of the British Comn;oriwealth, 78, 577. Grennert, L., Perrson, P.-H., Gennser, G., Kullander, S., acd Thorell, J. (1 976) : Lancet, 1, 4. Horne. C. H. W., Towler, C . M., and Milne, G. D. (1 974) : Journal of Clinical Pathology, (in press) Jandial, V., Campbell, D. M., Towler, C. M., McGillivray, I., and Horne, C. H. W. (1976): British Journal of Obstetrics and Gynaecology, (in press). Radiochemical Centre (1973) : HPL Immunoassay Kit. Singer, W., Desjardins, P., and Friesen, H. G. (1970): Obstetrics and Gynecology, 36, 223. Tatra, G., Breitenecker, G., and Gruber, W. (1974): Archiv fur Cynakologie, 217, 383. Tatra, G., Placheta, P., and Breitenecker, G . (1975): Wiener klinische Wochenschrqft, 87, 279. Thompson, A., Horne, C. H. W., Steel, H., and Goudie, R. B. (1969): Nature, London, 221, 289. Towler, C. M., Jandial, V., Horne, C. H. W., and Bohn, H. (1976): British Journal of Obstetrics and Gynaecology, 83, 368.