- Email: [email protected]
Alpha fetoprotein in maternal serum: A new marker for detection of fetal distress and intrauterine death
Alpha fetoprotein in maternal serum: A new marker for detection of fetal distress and intrauterine death MARKKU
SEPPIILIC,
ERKKI Helsinki,
M.D.*
RUOSLAHTI,
M.D.
Finland
Increase in circulating maternal alpha fetoprotein (AFP) is associated with intrauterine fetal death. The AFP levels in 13 women whose fetuses died in utero were from 280 to 9,000 ng. per milliliter. Nine of them (70 per cent) were above 530 ng. per milliliter, which was the upper normal level in 6.5 uncomplicated pregnancies. In high-risk pregnancies, the AFP test correctly predicted 60 per cent of the cases with fetal distress and 92 per cent of the cases with normal fetoplacental function. If maternal AFP levels were above 800 ng. per milliliter, fetal distress or intrauterine fetal death occurred in 85 per cent of the cases, and intrauterine fetal death occurred in all 6 cases where the maternal AFP concentration was higher than 1,075 ng. per milliliter. The increase in maternal AFP took place before the fetal death. Clinically established fetal distress was correctly predicted by the AFP test in 2 women whose estriol excretion was normal. The results suggest that the AFP test contributes to the biochemical detection of fetal distress and intrauterine death.
T H E D I A G N 0 S I S Of intraUterine fetal distress represents a major problem in obstetrics. Among biochemical tests, determination of urinary estriol has been widely used in clinical practice to detect fetoplacental dysfunction. Comparison of five methods for monitoring fetal distress indicated that the best predictive efficacy could be obtained if simultaneous determinations of estriol and circulating human placental lactogen (HPL) were used.= Even in otherwise normal pregFrom the Department II of Obstetrics and Gynecology, University Central Hospital, and the Department of Serology and Bacteriology, University of Helsinki. Supported by grants from the Sigrid Iuse’lius Foundation and the Finnish Medical Research Council. Received
for
f;;;pted
for publication
publication
July August
II,
1972. II,
*Present address: Department II of Obstetrics and Gynecology, University Central Hospital, Helsinki, Finland.
nancies, complications may be predicted by serial estimations of circulating HPL.2 Alpha fetoprotein (AFP) occurs at high serum concentrations during intrauterine life.3 AFP is synthesized by the fetal liver4 and yolk sac.5 It has recently become evident that small amounts of AFP are present in the sera of healthy adults.G The circulating maternal AFP increases during pregnancy and is usually highest during the third trimester before term.’ Wes used radioimmunoassay to determine maternal serum AFP levels in normal and high-risk pregnancies and found that the AFP test is useful in predicting fetal distress and intrauterine death. Materials
and
methods
Two hundred and six serum samples from 143 pregnant women in the second half of pregnancy were investigated. Sixty-five women had normal pregnancy, and 78 pregnancies were pathologic. Living infants were
Alpha
fetoprotein
in maternal
1000
serum
49
+
500
+ + + l
8 .
----$---9 1 aP z!. 2 8i
___^- -- ----
l +
t t 7
Normal
Fig. 1. Highest observed The dashed line indicates once.
? . . l ”
l
”.
Toxemia
”
or
Liver
individual AFP levels in 143 women the upper normal level. The value
born in 65 of these pregnancies, including 44 women with hypertension, toxemia, or pre-eclampsia and 21 women with a liver disorder. Intrauterine fetal death occurred in 13 cases, where hypertension or toxemia was present in 6 cases and liver disorder in 2. There was no obvious cause for the fetal death in 5 cases. Sixty-nine of the 78 high-risk pregnancies could be classified into the group with signs of severe fetoplacental dysfunction (20 cases) or those with normal fetoplacental function’ (49 cases). Fetal distress was assumed when two or more of the following signs were present: abnormal fetal heart
____-^_---+ ++
!
t Q
l
+
+
- - - - -;- ^- -
l .
+
+
2
in the second half of each individual
of pregnancy. is given only
action (late deceleration or variation higher than 120 to 160 beats per minute), fetal acidosis below pH 7.25, meconium in the amniotic fluid, retarded intrauterine fetal growth, and intrauterine fetal death. In this group, results of the AFP test were classified as higher (positive) or lower (false negative) than the upper normal AFP level in pregnancy (mean i- 2 x standard deviation) . Normal fetoplacental function was assumed when fetal heart action, pH values in the fetal blood, Apgar score (7 or more). birth weight, amniotic fluid, and the macroscopic appearance of the placenta were normal. In this group, the results of the AFP
50
Sepp&
and
January 1, 1973 Am. J. Obstet. Gynecol.
Ruoslahti
1000
CB
500
(B
e e %I 8 @
. __---!
-----
_-------_-
..
0 : *
0 4 VW..-- I----t” . .
; l
. . . 0
:
. .
. I
i @ . ----‘f---?B .. t i
l
.I. 1 .
.
.:. .. 29 :
..9. . .
l
.
l
0 . .
26
-30
Pregnancy
Fig. 2. Maternal fetoplacental level.
serum AFP levels distress. + = Intrauterine
.
-r
. 21-25
U.
.
.
31-35
36-40
week
in high-risk pregnancies. Circles fetal death. The dashed line
test were classified as normal or false positive (higher than the upper normal level). AFP levels of 400, 530, 800, and 1,000 ng. per milliliter were chosen to investigate the efficacy of the AFP test in predicting fetal distress. Serum AFP concentrations were measured by radioimmunoassay.* The samples were tested at 1 :50 or sometimes higher dilutions up to 1:5,000. Results Maternal serum AFP levels varied from 53 to 550 ng. per milliliter in 65 uncomplicated pregnancies. The upper normal level was 530 ng. per milliliter (Fig. 1). Seven
*
indicate indicates
cases with severe the upper normal
of 44 women (16 per cent) in the toxemia group had AFP levels higher than 530 ng. per milliliter, and 2 of 21 women with a liver disorder (9.5 per cent) exceeded this value. In high-risk pregnancies where live-born infants were delivered, 14 per cent of the mothers had AFP concentrations above the upper normal level. The AFP concentrations in 13 women with intrauterine fetal death varied from 280 to 9,000 ng. per milliliter (Fig. 2). Increase in the maternal AFP level took place before the fetal death occurred, and the highest level was found 3 days after death while the fetus was still in utero (Fig. 3). Nine of 13 women ( 70
Alpha
10
2, W**hti
Fig. 3. Circulating
22 of
13
21
25
in maternal
serum
51
26
pr.gn*ncy
maternal AFP in a representative
per cent) whose fetuses died in utero had AFP levels higher than the upper normal level, Fetal death occurred in all cases where AFP levels were higher than 1,075 ng. per milliliter (6 cases) . Among 78 high-risk pregnancies, severe fetoplacental dysfunction was established in 20 and normal fetoplacental function in 49 cases. In fetal distress, AFP levels above 530 ng. per milliliter occurred in 60 per cent. Two such women had normal urinary estriol excretion. In the group with normal fetoplacental function, the AFP test at the 530 ng. per milliliter level gave a correct result in 92 per cent and a false positive result in 8 per cent of the cases (Table I). The best predictive efficacy of fetal distress by the AFP test was achieved at the 400 ng. per milliliter level, where fetal distress was correctly indicated in 75 per cent of the cases and normal fetoplacPnta1 function in 80 per cent of the cases. In 13 high-risk pregnancies where circulating maternal AFP levels were higher than 800 ng, per milliliter, the fetus was
fetoprotein
case where the fetus died in utero.
Table I. Efficacy of the AFP test for early detection of fetooplacental dysfunction in 69 high-risk pregnancies evere fetoplacental dysfunction (20)
AFP
‘mi)
I
(%I
>
400
75
> >
530” 800
60 55 40
1 ma0
/
(%) 25 40 45 60
Normal fetoplacental funcliofl (49)
I
i%)
96 96
I
(%.J 20 8 4 4
*uppernormal Irvrl. either severely distressed or dead cent of the cases (Fig. 2).
in 85 per
Comment The origin of elevated AFP during pregnancy may be the fetus. This has been suggested by a significant correlation between the newborn and maternal AFP levels and by the fact that the newborn level is 300 to 600 times higher than the maternal level at term.? Another possibility is that pregnant
52
Seppiilti
and
Ruoslahti
women themselves produce AFP more than nonpregnant individuals. Presence of a genetic marker in AFP would permit settling of this question, but polymorphism has not so far been detected in AFP. The maternal peak level of AFP occurs around the thirty-second week of pregnancy, where the fetal production of AFP begins to decrease.3 Decreasing AFP concentrations in fetal serum between 13 and 22 weeks’ gestation are attributable to an increase in the rate of growth in the conceptus that exceeds the increase in the amount of AFP synthesized during this period. Between 22 and 32 weeks’ gestation, the relative amount of circulating AFP in the fetus remains steady.” An increase above the normally occurring pregnancy level of AFP was found 10 days before intrauterine fetal death in a representative case (Fig. 3). The time interval between the elevation of AFP and demonstration of fetal distress varied from 1 to 14 days. Because the samples were mostly taken at weekly intervals, the relationship between the onset of fetal distress and elevation of maternal AFP could not be accurately established by the present data. No clinical decisions were based on the AFP determinations in the present series. The increase in AFP was similar in women with toxemia and liver disorder. Elevated maternal AFP levels in fetal distress may result from either an increased production of AFP by the distressed fetus or placental leakage of fetal elements. A transamniotic route is also possible. While all these mechanisms may be operating, there is preliminary evidence that distressed newborn infants may have higher AFP levels than normal infants.” In the presence of fetal distress, the AFP REFERENCES
1. Keller, P. J., Baertschi, LJ., Bader, P., et al.: Lancet 2: 729, 1971. 2. Letchworth, A. T., and Chard, T.: Lancet 1: 704, 1972. 3. Gitlin, D., and Boesman, M.: J. Clin. Invest. 45: 1826, 1966. 4. Gitlin, D., and Boesman, M.: J. Clin. Invest. 46: 1010, 1967.
January 1, 1973 Am. J, Obstet. Gynecol.
test gave a correct result in 60 per cent of the cases at the 530 ng. per milliliter level and in 75 per cent of the cases at the 400 ng. per milliliter level. In a previous study where similar criteria for fetal distress were used,l the predictive efficacy of urinary estriol excretion was 69 per cent and that of HPL was 100 per cent. In high-risk pregnancies where the fetoplacental function was subsequently proved normal, the predictive efficacy of the AFP test (92 per cent) was higher than that previously shown for either estriol (88 per cent) or HPL (65 per cent) .I Calculated from the data given in Fig. 2, it appears that one may safely wait for a rise of AFP to 400 ng. per milliliter in 93 per cent and to 530 ng. per milliliter in 92 per cent of the cases before intervening in highrisk pregnancies. Fetal distress was correctly indicated by the AFP test in 2 cases at a time when estriol determinations showed normal excretion. This is the first introduction of the AFP test in the biochemical monitoring of the fetus in fetoplacental distress. Although preliminary, our results suggest that determination of maternal AFP provides additional information on fetal well-being in utero. Radioimmunoassay is a standard method in many clinical laboratories, and reagents for radioimmunoassay of AFP are available from at least one commercial source. The upper normal AFP level during the third trimester is so high that it may even be detectable by methods other than radioimmunoassay.
We thank Miss Sirkka Soikkeli, Miss Hannele Kallio, Mrs. Marjatta Vuori, and Miss Leena Salminen for technical assistance. 5. Gitlin, D., and Pericelli, A.: Nature 228: 995, 1970. 6. Ruoslahti, E., and SeppZ, M.: Nature 235: 161, 1972. 7. SeppHlL, M., and Ruoslahti, E.: AM. J. OBSTET. GYNECOL. 112: 208, 1972. 8. Ruoslahti, E.> and SeppllZ, M.: Int. J. Cancer 8: 374, 1971. 9. Nsrgaard-Pedersen, B.: Personal communication 1.
SEPPIILIC,
ERKKI Helsinki,
M.D.*
RUOSLAHTI,
M.D.
Finland
Increase in circulating maternal alpha fetoprotein (AFP) is associated with intrauterine fetal death. The AFP levels in 13 women whose fetuses died in utero were from 280 to 9,000 ng. per milliliter. Nine of them (70 per cent) were above 530 ng. per milliliter, which was the upper normal level in 6.5 uncomplicated pregnancies. In high-risk pregnancies, the AFP test correctly predicted 60 per cent of the cases with fetal distress and 92 per cent of the cases with normal fetoplacental function. If maternal AFP levels were above 800 ng. per milliliter, fetal distress or intrauterine fetal death occurred in 85 per cent of the cases, and intrauterine fetal death occurred in all 6 cases where the maternal AFP concentration was higher than 1,075 ng. per milliliter. The increase in maternal AFP took place before the fetal death. Clinically established fetal distress was correctly predicted by the AFP test in 2 women whose estriol excretion was normal. The results suggest that the AFP test contributes to the biochemical detection of fetal distress and intrauterine death.
T H E D I A G N 0 S I S Of intraUterine fetal distress represents a major problem in obstetrics. Among biochemical tests, determination of urinary estriol has been widely used in clinical practice to detect fetoplacental dysfunction. Comparison of five methods for monitoring fetal distress indicated that the best predictive efficacy could be obtained if simultaneous determinations of estriol and circulating human placental lactogen (HPL) were used.= Even in otherwise normal pregFrom the Department II of Obstetrics and Gynecology, University Central Hospital, and the Department of Serology and Bacteriology, University of Helsinki. Supported by grants from the Sigrid Iuse’lius Foundation and the Finnish Medical Research Council. Received
for
f;;;pted
for publication
publication
July August
II,
1972. II,
*Present address: Department II of Obstetrics and Gynecology, University Central Hospital, Helsinki, Finland.
nancies, complications may be predicted by serial estimations of circulating HPL.2 Alpha fetoprotein (AFP) occurs at high serum concentrations during intrauterine life.3 AFP is synthesized by the fetal liver4 and yolk sac.5 It has recently become evident that small amounts of AFP are present in the sera of healthy adults.G The circulating maternal AFP increases during pregnancy and is usually highest during the third trimester before term.’ Wes used radioimmunoassay to determine maternal serum AFP levels in normal and high-risk pregnancies and found that the AFP test is useful in predicting fetal distress and intrauterine death. Materials
and
methods
Two hundred and six serum samples from 143 pregnant women in the second half of pregnancy were investigated. Sixty-five women had normal pregnancy, and 78 pregnancies were pathologic. Living infants were
Alpha
fetoprotein
in maternal
1000
serum
49
+
500
+ + + l
8 .
----$---9 1 aP z!. 2 8i
___^- -- ----
l +
t t 7
Normal
Fig. 1. Highest observed The dashed line indicates once.
? . . l ”
l
”.
Toxemia
”
or
Liver
individual AFP levels in 143 women the upper normal level. The value
born in 65 of these pregnancies, including 44 women with hypertension, toxemia, or pre-eclampsia and 21 women with a liver disorder. Intrauterine fetal death occurred in 13 cases, where hypertension or toxemia was present in 6 cases and liver disorder in 2. There was no obvious cause for the fetal death in 5 cases. Sixty-nine of the 78 high-risk pregnancies could be classified into the group with signs of severe fetoplacental dysfunction (20 cases) or those with normal fetoplacental function’ (49 cases). Fetal distress was assumed when two or more of the following signs were present: abnormal fetal heart
____-^_---+ ++
!
t Q
l
+
+
- - - - -;- ^- -
l .
+
+
2
in the second half of each individual
of pregnancy. is given only
action (late deceleration or variation higher than 120 to 160 beats per minute), fetal acidosis below pH 7.25, meconium in the amniotic fluid, retarded intrauterine fetal growth, and intrauterine fetal death. In this group, results of the AFP test were classified as higher (positive) or lower (false negative) than the upper normal AFP level in pregnancy (mean i- 2 x standard deviation) . Normal fetoplacental function was assumed when fetal heart action, pH values in the fetal blood, Apgar score (7 or more). birth weight, amniotic fluid, and the macroscopic appearance of the placenta were normal. In this group, the results of the AFP
50
Sepp&
and
January 1, 1973 Am. J. Obstet. Gynecol.
Ruoslahti
1000
CB
500
(B
e e %I 8 @
. __---!
-----
_-------_-
..
0 : *
0 4 VW..-- I----t” . .
; l
. . . 0
:
. .
. I
i @ . ----‘f---?B .. t i
l
.I. 1 .
.
.:. .. 29 :
..9. . .
l
.
l
0 . .
26
-30
Pregnancy
Fig. 2. Maternal fetoplacental level.
serum AFP levels distress. + = Intrauterine
.
-r
. 21-25
U.
.
.
31-35
36-40
week
in high-risk pregnancies. Circles fetal death. The dashed line
test were classified as normal or false positive (higher than the upper normal level). AFP levels of 400, 530, 800, and 1,000 ng. per milliliter were chosen to investigate the efficacy of the AFP test in predicting fetal distress. Serum AFP concentrations were measured by radioimmunoassay.* The samples were tested at 1 :50 or sometimes higher dilutions up to 1:5,000. Results Maternal serum AFP levels varied from 53 to 550 ng. per milliliter in 65 uncomplicated pregnancies. The upper normal level was 530 ng. per milliliter (Fig. 1). Seven
*
indicate indicates
cases with severe the upper normal
of 44 women (16 per cent) in the toxemia group had AFP levels higher than 530 ng. per milliliter, and 2 of 21 women with a liver disorder (9.5 per cent) exceeded this value. In high-risk pregnancies where live-born infants were delivered, 14 per cent of the mothers had AFP concentrations above the upper normal level. The AFP concentrations in 13 women with intrauterine fetal death varied from 280 to 9,000 ng. per milliliter (Fig. 2). Increase in the maternal AFP level took place before the fetal death occurred, and the highest level was found 3 days after death while the fetus was still in utero (Fig. 3). Nine of 13 women ( 70
Alpha
10
2, W**hti
Fig. 3. Circulating
22 of
13
21
25
in maternal
serum
51
26
pr.gn*ncy
maternal AFP in a representative
per cent) whose fetuses died in utero had AFP levels higher than the upper normal level, Fetal death occurred in all cases where AFP levels were higher than 1,075 ng. per milliliter (6 cases) . Among 78 high-risk pregnancies, severe fetoplacental dysfunction was established in 20 and normal fetoplacental function in 49 cases. In fetal distress, AFP levels above 530 ng. per milliliter occurred in 60 per cent. Two such women had normal urinary estriol excretion. In the group with normal fetoplacental function, the AFP test at the 530 ng. per milliliter level gave a correct result in 92 per cent and a false positive result in 8 per cent of the cases (Table I). The best predictive efficacy of fetal distress by the AFP test was achieved at the 400 ng. per milliliter level, where fetal distress was correctly indicated in 75 per cent of the cases and normal fetoplacPnta1 function in 80 per cent of the cases. In 13 high-risk pregnancies where circulating maternal AFP levels were higher than 800 ng, per milliliter, the fetus was
fetoprotein
case where the fetus died in utero.
Table I. Efficacy of the AFP test for early detection of fetooplacental dysfunction in 69 high-risk pregnancies evere fetoplacental dysfunction (20)
AFP
‘mi)
I
(%I
>
400
75
> >
530” 800
60 55 40
1 ma0
/
(%) 25 40 45 60
Normal fetoplacental funcliofl (49)
I
i%)
96 96
I
(%.J 20 8 4 4
*uppernormal Irvrl. either severely distressed or dead cent of the cases (Fig. 2).
in 85 per
Comment The origin of elevated AFP during pregnancy may be the fetus. This has been suggested by a significant correlation between the newborn and maternal AFP levels and by the fact that the newborn level is 300 to 600 times higher than the maternal level at term.? Another possibility is that pregnant
52
Seppiilti
and
Ruoslahti
women themselves produce AFP more than nonpregnant individuals. Presence of a genetic marker in AFP would permit settling of this question, but polymorphism has not so far been detected in AFP. The maternal peak level of AFP occurs around the thirty-second week of pregnancy, where the fetal production of AFP begins to decrease.3 Decreasing AFP concentrations in fetal serum between 13 and 22 weeks’ gestation are attributable to an increase in the rate of growth in the conceptus that exceeds the increase in the amount of AFP synthesized during this period. Between 22 and 32 weeks’ gestation, the relative amount of circulating AFP in the fetus remains steady.” An increase above the normally occurring pregnancy level of AFP was found 10 days before intrauterine fetal death in a representative case (Fig. 3). The time interval between the elevation of AFP and demonstration of fetal distress varied from 1 to 14 days. Because the samples were mostly taken at weekly intervals, the relationship between the onset of fetal distress and elevation of maternal AFP could not be accurately established by the present data. No clinical decisions were based on the AFP determinations in the present series. The increase in AFP was similar in women with toxemia and liver disorder. Elevated maternal AFP levels in fetal distress may result from either an increased production of AFP by the distressed fetus or placental leakage of fetal elements. A transamniotic route is also possible. While all these mechanisms may be operating, there is preliminary evidence that distressed newborn infants may have higher AFP levels than normal infants.” In the presence of fetal distress, the AFP REFERENCES
1. Keller, P. J., Baertschi, LJ., Bader, P., et al.: Lancet 2: 729, 1971. 2. Letchworth, A. T., and Chard, T.: Lancet 1: 704, 1972. 3. Gitlin, D., and Boesman, M.: J. Clin. Invest. 45: 1826, 1966. 4. Gitlin, D., and Boesman, M.: J. Clin. Invest. 46: 1010, 1967.
January 1, 1973 Am. J, Obstet. Gynecol.
test gave a correct result in 60 per cent of the cases at the 530 ng. per milliliter level and in 75 per cent of the cases at the 400 ng. per milliliter level. In a previous study where similar criteria for fetal distress were used,l the predictive efficacy of urinary estriol excretion was 69 per cent and that of HPL was 100 per cent. In high-risk pregnancies where the fetoplacental function was subsequently proved normal, the predictive efficacy of the AFP test (92 per cent) was higher than that previously shown for either estriol (88 per cent) or HPL (65 per cent) .I Calculated from the data given in Fig. 2, it appears that one may safely wait for a rise of AFP to 400 ng. per milliliter in 93 per cent and to 530 ng. per milliliter in 92 per cent of the cases before intervening in highrisk pregnancies. Fetal distress was correctly indicated by the AFP test in 2 cases at a time when estriol determinations showed normal excretion. This is the first introduction of the AFP test in the biochemical monitoring of the fetus in fetoplacental distress. Although preliminary, our results suggest that determination of maternal AFP provides additional information on fetal well-being in utero. Radioimmunoassay is a standard method in many clinical laboratories, and reagents for radioimmunoassay of AFP are available from at least one commercial source. The upper normal AFP level during the third trimester is so high that it may even be detectable by methods other than radioimmunoassay.
We thank Miss Sirkka Soikkeli, Miss Hannele Kallio, Mrs. Marjatta Vuori, and Miss Leena Salminen for technical assistance. 5. Gitlin, D., and Pericelli, A.: Nature 228: 995, 1970. 6. Ruoslahti, E., and SeppZ, M.: Nature 235: 161, 1972. 7. SeppHlL, M., and Ruoslahti, E.: AM. J. OBSTET. GYNECOL. 112: 208, 1972. 8. Ruoslahti, E.> and SeppllZ, M.: Int. J. Cancer 8: 374, 1971. 9. Nsrgaard-Pedersen, B.: Personal communication 1.