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Serum unconjugated estriol levels in the third trimester and their relationship to gestational age
Serum unconjugated estriol levels in the third trimester and their relationship to gestational age JOHN
E.
JOSEPH ALLEN
BUSTER,
M.D.*
SAKAKINI.
JH..
P.
KlLL.Ahl.
WILLIAM
El Paw,
H.
M.D
51.1).
SC:RAG(;,
iU.1)
Teaas
Maternal unconjugated estriol levels ~LWW measured throughout the 28 to 41 week intenlal in two groups of accuratrly dated normal pregnancies. The first group consisted of randomly sampled pregnancies on which 285 unconjugated estriol determinations were prrformed. The logarithms of the mean ualues plotted into a positive sloping, relatively .straight line ulhich uus disrupted bl; a plateau orig-inating at 31 to 32 weeb and terminated at 35 weeks xlhrrc therr began a .steep surge to a point at 36 weeks (SUrgQ point) that returned value5 to fit the previously established .rtraight line. To Avestigate thesejindings in individual pregnancies, a second group nine subjects was studied with .serial unconjugated e.ytriol determinations. In all nine of these subjects, the Aurge point could be identified statistically and occurred at a mean gestational age C{ 35.0 + 0.6 (1 S.D.) weeks. Data,from the first group oJ mndomlp sampled pregnancies indica.te that the .curge point occurred around a mean gPvtationa1 aie of 36.0 weeks and was conJirmtd by data from the second group .srrially sampled individual subjects showirq th,r surge point as a statistically dejnable marker in normal pregnancies.
(rf
of
I N NORMAL pregnancy a continued increase in the rate of fetal adrenal growth occurs between 28 and 41 weeks’ gestation and becomes particularly marked between 34 and 36 weeks.’ This is associated with a sharp rise in circulating fetal and amniotic fluid cortisol levels.‘, ’ The exposure of fetal tissues to rising cortisol levels is probably the initiating event in a series of final maturation steps that include increasing pulmonary surfactant production and rising glycogen stores.“. ’ The mechanisms controlling fetal adrenal growth and steroidogenesis are largely modulated by the fetal hypothalamus and pituitary. ‘-” Accelerated fetal adrenal growth occurring during the 34 to 36 week interval has been linked to a concomitant rise in levels of circulating fetal pituitary prolartin.” In addition, rising cortisol levels are associated with a simultaneous
From the Deportment oj Olxtrtnc~ William Aeaumont Army Medical Rmived
Accepted
for
publicnlwn
December-
Suptembrl-
fall (suppression) in circulating fetal pituitary ACTH,‘O implying, as has been demonstrated in sheep, that the fetal adrenal (probably definitive cortex) has become increasingly sensitive to ACTH. ” Unconjugated estriol levels. as measured in the peripheral maternal circulation, are known to correlate closely with fetal adrenal steroidogenesis and the fetal corticotrophic mechanisms which regulate it. 7. ‘. I2 This consideration led us to study maternal unconjugated estriol levels in a series of uncomplicat.ed pregnancies at carefully documented intervals between 28 and 41 weeks to observe if known changes in fetal adrenal steroidogenesis would be reflected by alterations in maternal serum estriol concentrations.
Materials and methods Serum unconjugated estriol was measured in a series of normal, carefully dated pregnancies by a specific, rapid, nonchromatographic radioimmunoassay described previously.‘3 Serum was obtained by peripheral venipuncture and stored after separation at -20” C. until assayed. Each series of samples was run in the same assay such that between-sample variation was held to < 5.0 per cent. Blood samples were obtained between 13:30 and 15:30 hours from ambulatory outpatients with uncomplicated pregnancies delivering
arid Gywx~log~, Centw.
9. 1975.
16, 1975.
Reprint wqursts: .411rn P. Rillam, M.D., Department Obstetrics and Gynecology, William Beaumont Army Medical Center, El Pasn, TPXCIS 79920. Pwwnt addww: John Rcjproductive Biology, Canon St., Torm~r,
E. Buster, M.D., Diwsion Harbor General Hospital, Ccllifornin 90509.
oj 1000
oJ
K’.
672
Volume Number
Third-trimester
125 5
I
I
I
28
30
I 32
GESTATIONAL
34
)
)
I
,
36
38
40
42
AGE
serum unconjugated
estriol
673
(WEEKS)
Fig. 1. The means and standard errors for the means from 285 unconjugated estriol determinations between weeks 28 and 41 of gestational age on a semilogarithmic plot. Figures in parentheses indicate the numbers of samples for each point.
healthy term neonates. Pregnancies were dated on the basis of accurate and regular menstrual histories, confirmed by accurate midtrimester sizes, and reconfirmed by the pediatrician’s examination utilizing standardized published weight, physical, and neurologic criteria. i4-r6 Two groups of pregnant women were studied. The first group consisted of randomly selected subjects from whom one to nine samples were obtained between 28 and 41 weeks’ gestation, The great majority were sampled from one to three times for a total of 285 samples. The second group consisted of nine subjects who provided serial samples. Clinical data for these subjects are summarized in Table I. Data obtained from the first group of subjects were treated in the following manner. The means and standard error of the means from the 285 samples were placed on a logarithmic ordinate plotted against gestational age on a linear abscissa (Fig. 1). The logarithmic transformed mean values plotted into a positive sloping, nearly straight line disrupted by a plateau originating at the 31 to 32 week point and terminating at 35 weeks where a steep surge began and terminated at the 36 week point (surge point) that returned values to fit the previously established straight line. In order to confirm the above impression, a least-squares regression line with 95 per cent confidence limits was placed through all points between 28 and 40 weeks with deliberate exclusion of the 33 to 34 week and 35 week point (Fig. 2). The fit of this line was very close (r = 0.997). It was noted that the 33 to 34 week point (A) fell exactly on the 95 per cent confidence line and the 35 week(B) point well below it, indicating that probably neither point belonged to the straight line logarithmic transformed relationship described above. The 33 to 34 week point (A) and the 35 week point(B) were then compared by unpaired t tests with corresponding points predicted by the regression
Fig. 2. Logarithmic transformed regression line with 95 per cent confidence limits fitted to points between 28 and 40 weeks with deliberate exclusion of the points at 33 to 34 and 35 weeks where the numbers plateau from the trend of surrounding data points. The 33 to 34 week point (A), and the 35 week point (II), the 35 to 36 week interval (C), and the 41 week point (0) are tested statistically for both their relationship to the 95 per cent confidence intervals from the logarithmic transformed regression line and the corresponding values predicted from that line by unpaired t tests.
32282420 = 16. f 14p 12; IOB 6:, w 6-
$fSEY i--J-+ 28 30 32 14 36 38 40 42 GESTATIONAL
Fig. 3. Statistical regression line
AGE
(WEEKS)
definition of the surge point. A least-squares placed through the four points between 29.5
and 35 weeks and extrapolated
to week 36 point gives a
predicted future value ?95 per cent confidence first value falling outside of the 95 per cent interval is defined as the surge point.
limits. This confidence
line (Fig. 2). For the calculation, it was assumed that corresponding points had the same variance. The difference between points at the 33 to 34 week interval was not significant (p > 0.05) but the difference at 35 weeks was highly significant (p < 0.01). In order to investigate the significance of the surge point, the mean values and variances for the samples obtained to compute the 35 to 36 week points were compared by unpaired t tests, also confirming a highly significant difference (p > 0.01) (Fig. 2). The surge point was also defined another way (Fig. 3). A least-squares regression line was placed through the four points between 29.5 and 35 weeks and extended to week 36 with extrapolation of a future value at 36 weeks &95 per cent confidence limits of 11.8 -C 1.3 ng per milliliter. The measured level at 35 weeks of 15.3 ng. per milliliter fell
674
July 1, IY7ti Am. J. Obstet. Gynecol.
Buster et al.
L&o,
4imu 283032 343638 4042 :‘8 JL 24 1
GESTATIONAL
AGE (WEEKS)
Fig. 4. Individual plots troni nine pregnant sunjects with accurate dates. The surge point occurs III) earlier than 35.0 weeks and no later than 37.0 weeks with a inean of 86.0 2 0.6 (I SD.). In eight of nine subjects labor occurred between 39..i and 41.0 weeks with a tnean surge point to labor interval of 3.9 2 0.6 (1 SD.) weeks. In one subjec I (,J.I..), labor occurred at 3X.5 weeks with a surge point to labor interval of‘ 2.5 weeks.
well above the 95 per cent conhdence inter\-al predicted bv the line.” Data from the second group of subjects were treated in the following manner. Individual points were placed on a semilogarithmic plot exactly as described with the mean data from the first group. The surge point was then calculated and defined as follows (Figs. 3 and 4). With the use of the logarithms of the estriol concentrations from the first 3 weeks of observation a leastsquares regression line was calculated and plotted. The expected value and its upper 95 per cent confidence limit for the next week’s concentration were then calculated from the slope of the line. Whenever the actual observed serum concentration fell below this limit it was assumed to be a part of the previously calculated regression line and a new regression line was calculated incorporating this additional data point. The next actual concentration would then be compared to the predicted next upper 95 per cent confidence limit and the process was continued until the first measured concentration exceeded the upper 95 per cent conh-
dence limit. This point was considered as not part of this regression line and was defined as the surge point.
Results Data from the first group of pregnancies studied are shown in Fig. I. This figure depicts the logarithmic means and SXM. from 285 serum samples measured for unconjugated estriol levels plotted against the confirmed gestational age in weeks. The over-all positive logarithmic transformed linear slope and the transient downward displacement between 31 to 32 and 35 weeks occur as were noted previously. Although the point at 32 to 33 weeks is not significantly different (p > 0.05) from the corresponding value predicted by the regression line, this point falls on the lower 95 per cent confidence band for the entire line, implying that it probably does not belong to the straight logarithmic relationship described by the other points. The point at 33 to 34 weeks appears, therefore, to be transitional since it becomes significant if moved forward another O.:‘, weeks (p < 0.05).
Volume Number
Table
Third-trimester
125 5
I. Clinical
Subject J. C.
data from
nine pregnant
subjects
selected
for sequential
serum unconjugated
Gravida
PCWU
Fetal wt. at delivery (Gm.)
Gest. age at delivery (wk.)
27
4
1
2,948
41.0
9
Cord
40.5 39.5 39.5 40.0 40.0 40.0 40.0 38.5
9 10 9 9 10 9 10 9
None None None None None None None None
2.5
1
0
2,693
C. B. L. B. C. M. c. y. c. M’. G. R. J. L.
24 27 18 24 25 27 25
1 3 2 2 2 4 3
0 2 0 1 1
3,218 3,600 3,303 2,562 3,473 3,062 2,736
:
A second decline occurs at 41 weeks. This point falls below an extension of the 95 per cent confidence bands and is also significantly lower (p < 0.05) than the value predicted for 41 weeks by the regression line (Fig. 2). Fig. 3 depicts placement through the mean data points of‘ ;I least squares regression line as it is used to define rhe location of the surge point. Data f‘rom the group of nine individual pregnancies studied with serial samples is graphed on individual plots in Fig. 4 and tabulated in Table II. Fig. 4 depicts placements of regression lines, predicted values ?95 per cent confidence intervals, and the placement of the surge point as described. The mean gestational age at which the surge point occurred from all nine subjects was 36.0 rt 0.6 (I S.D.) weeks. The earliest placement was at 35.0 weeks and the latest was 37.0 weeks. In eight of nine subjects, spontaneous labor occurred between 39.0 ancl 40.5 weeks, and the mean surge point to labor interval was 3.9 -C 0.6 (1 SD.) weeks. In one of the nine subjects (J.L.), labor occurred at 38.5 weeks and was associated with a surge point to labor interval of 2.5 weeks.
Comment Numerous publications in discussing normal values for serum and urinary estriol during the third trimester have depicted a smooth curve which gradually swings upward until term.‘8-21 The 31 to 35 week plateau and 35 to 36 week surge described in this study have so far been unacknowledged. These phenomena may, in part, he unique to the unconjugated fraction of circulating estriol since this moiety is almost exclusively fetoplacental in origin and has not been shifted and metabolized through multiple maternal compartments as are the various estriol conjugates.“’ 23 Another explanation may lie with inaccurate dating of gestational ages in some series. Finally, in several publications the data has been condensed into 2 to 4 week intervals such that the rather short-term events described here would remain unnoticed.‘x-2’
APw
Complications
(5 min.)
compression during labor, otherwise none
Table II. Gestational age at time of surge surge point to labor interval Sd@
vi. C. L. C. C. c. G. J.
point
and
Gestational agg at surge point (wk.)
Surge point to labor interval (wk.)
37.0 37.0 35.5 35.0 35.5 36.0 36.0 36.0 36.0
4.0 3.0 3.5 4.0 4.5 3..5 4.0 4.5 2.5*
B. B. M. Q. w. R. L.
Mean ? 1 S.D.
675
studies
4-e
M. B.
estriol
36.0
f 0.6
3.9 t 0.6
*Labor at 38.5 weeks and not included in mean calculation. Any attempt to explain the downward deviations from an otherwise positive sloping logarithmic transformed line must be speculative with the present state of knowledge. It is plausible that these effects are due to increasing levels of fetal cortisol leading to decreased levels of ACTH.‘s ‘* “This in turn may cause decreased production of fetal dehydroepiandrosterone sulfate (DHEA-S) with a resultant decrease in placental estriol production. 7. 8 The 35 to 36 week estriol surge is more difficult to explain. The resumption of the previously established upswinging logarithmic transformed linear relationship may represent a sensitizing phenomenon such that the fetal cortex becomes more responsive to fetal ACTH and continues to produce increasing amounts of DHEA-S despite decreased circulating ACTH. Since circulating fetal prolactin rises sharply after week 34 and since it is believed to enhance fetal adrenal growth, lipid biosynthesis, and cholesterol (steroid precursor) storage, ‘it is possible that prolactin may promote increasing fetal adrenal zone sensitivity to ACTH through a permissive or additive effect. A similar situation may occur in the 40 to 41 week interval where the estriol curve slopes downward. The occurrence may reflect on further increases in fetal cortisol production and additional feedback ACTH sup-
676
Buster et al.
pression just prior to the onset of term labor.‘O In premature labor, an event associated with rising maternal estradiol levels and falling progesterone levels,” estrio1 concentrations have not been studied. However. the changes observed in estriol concentratiom at around 36 weeks may be more related to the steriod mediated maturation process active at that time rather than to the mechanisms triggering the onset of labor, as suggested by the findings in subject J. L.. who labored at 38.5 weeks, 2.5 weeks after the estriol surge poim. Determination of the surge point in individual pregnancies could be utilized as a noninvasive clinical test for dealing with patients \vho at-r first etKountet.ed during the third trirnesLe?- ~rntt drc corrrplicatrd b\ itiaccuraW or misleading riates. I‘he multitude of as well as frankI\ harartlous, probletns irritating, associated with “bad date” pregnancies are lvrll knowrt to all practicing obst,etricians. These data ittdicatc that. in il normal pregnatic v deVitr~d to deliver ,lt tcrnt. deterrninatior~ of tltrx surge poittt ~\ould permit: (1) an accurate c~stimation of’ the tcrrrt ~~;it‘tUriticJIl tlatc Lvithout other clinical irifortrla~iotl ;%II~ (2) intlicate thv
time at which adrenal steroidogenesis has ah.eaclv been shifted tol\xrd increasing corticoid produc tiorr and. 1~1 inference, increasitrg pulnionar\~ sttrfactatit produ(.tion. This latter point will cleat-l! require further, docurnentatioti. In order to utilize the surge poitil detet minatioti lorclinical purposes, it is essential that all sartiples be run through thr sa~ne assal in adjacent tubes so as to minitnizc rnrthodologir errors. It is, therefore. ot~t‘ \veekly blood samples f‘roln current prxtic-e to “bank” patients with poor dates with the expectation (4 running the entire series if. the need should arise. ‘The c;tlculation is best carried out with fout. or ntot-r points itt lvhich w plate a regression line. If three point> ;ttx‘ utilized. the regression line must fit WY\ closel\ in order to predic I ;I future value v;it tt ;I \vorkablv nat-roe !?5 per cent c,onfidence interval. Prelimittarv c-lirtical experience \\ith this approach has been cncx)uragirtg ‘I‘he authotx wish to thank 1)x.. Martin Vusynowity oi William Reautnont Army Medical Ctrnter ;uld Dr. John Rlarshall of Harbor General Hospital for their statistical assisraiicr ;iricl et~t‘o~irageiricttt.
REFERENCES
I, Spector,
W. S.: Handbook
of Biological
Data.
Philadel-
phia. 1956, W. B. Saunders Companv. 1’. ?153. 2. Murphy, B. E. P.: Does the human fetal adrenal plak a role in parturition? AM. J. OBSTEI.. GYNECOI.. 115: 521,
13.
11.
1973.
3. Montserrat, F. I)., and Tulchinsk\. D.: .I otal cortisol in amniotic Huid and fetal lung maturation, N. Engl. J. Med. 4.
.i. 6. 7.
8.
9.
10.
Il. 12.
292: 133, 1975. Liggens, G. C.: Endocrinology of the foetal maternal urrii. iti Shearman. R. P., editor: Human Reproductive Physiology, Oxford, 1972, Blackwell Scientific Puhlications. p. 138. Gluck. L.: Pulmonar! surfactant and neonatal respiratot\ distress. Hosp. Pratt. 6: 45, 1971. Diczfalusy. E.: Endocrine functions of rhe human fetus and placenta. AM. J. OBSTET. G~NECOL. 119: 419, 1974. Simmer, H. H., ‘Tulchinsky. D.. Gold, E. XI.. Frankland, 11.. Greidel, 51.. and Gold, .4. S.: On regulation of estrogen production hy cortisol and ACTH in human pregnancy at term, AM. J. OBSTET. GymcoL. 119: 283, 1974. Simmer. H. H.. Frankland, 11.. and Greidel, hf.: On the regulation of fetal and maternal 16~hydroxydehydroepiandrosterone and its sulfate by cortisol and ACTH in human pregnancy at term, AM. J. OBSTET. GYKECOL. 121: 646. 1975. Winters, A. J.. Colston, C., MacDonald. P. C.. and Porter, J. C.: Fetal plasma prolactin levels, J. Clin. Endocrinol. Metab. 41: 626. 1975. Winters. A. J., Oliver, C., C&ton. C., .ZZacDonald, P. C., and Porter, J, C.: Plasma ACTH in the human fetus and neonate as related to age and parturition, J. Clin. Endocrinol. Metab. 39: 269, 1974. Liggins, G. C.: Premature delivery of fetal iambs infused with glucocorticoids, J. Endocrinol. 45: 515. 1969. Whitt, G. G., Buster, J. E.. Killam, il. P., and Scragg, W. H.: Comparison of two giucocorticoid regimens for the acceleration of fetal lung maturation in premature labor. AM. J. OBSTET. GYNECOL. 124: 479. 1976.
15.
6. 7. 1 8.
9.
20.
21.
22.
23.
Abraham, C. b:.. Buster-, J, E., Samqjiik. L,. (&La, R.. ,rnti Hillman, B.: (:omparison between four radioimmunoaasays for plasma estriol, Anaiyt. Letters 7: 1 19, 1974. Lubchenco, I.. 0.: Assessment of gebtational age ,mti development at birth, Pediatr. Clin. North Am.17:3: 125. 1970. Usher. R., %lcLean. E‘.. and Scott. K. E.: Judgemenr 01 fetal age. II. Clinical significance of gestational age and an objective method for its assessment, Pediatr. Chin. North ,4m. 13: 835. 1966. Amiei-~I‘ison. t:.: Neurological evalualiotl of the matwit! of newborn infants, Arch. Dis. Child. 43: X9. 1968. Natreila, M. G.: Experimental statistics, National Bureau of Standards Handbook 91: 5, 1963. Clear!;. R. E.. and Young, P. C. Xl.: Serum unconjugated estriol in normal and abnormal pregnant!‘, .4\1. J. Ossref. GYNECOL. 118: 18. 1974. Lindberg. B. S.. Johansson. E. D. B.. and Niissen. B. A.: Plasma levels of nonconjugated estrone, estradiol-17 B. and estriol during uncomplicated pregnancy. dicta CIbstet. Gynecol. Stand. 32: (Suppl.) 21, 1971. Tulchinskv. I).. Hobel. C. H., and Korenman, S.: Radioligand assay for plasma unconjugated estriol. Ahf. J. ORSTET. GYNECOL. 111: 311. 1971. Scommegna, A.: Clinical uses of estriol assays, 2)1 Wlnn. R. M.. editor: Obstetrics and Gynecology Annual, New York, 1973, Appleton-Century-Crofts, p. 465. Kirdani. R. Y., Sampson, D.. AUurphy, G. P.. and Sandberg, A. A.: Studies on phenolic steroids in human subjects. XVI. Role of the kidney in the disposition of estriol, J. Chin. Endocrinoi. Metab. 34: 546. 1972. Diczfalusy. E., and Levitz, M.: Formation. metabolisln.
and transport
24.
of estrogen conjugates, in Bernstein,
S..
and Solomon. S.. editors: Chemical and Biological Aspects of Steroid Conjugation, New York, 1970. Springer Veriag, p. 291. Tamby Raja, R. I,.. Anderson, A. B. M., and ‘I‘urnbull, .4. C:.: Endocrine changes in premature iabour, Br. Med. ,J. 4: 67. 1974.
E.
JOSEPH ALLEN
BUSTER,
M.D.*
SAKAKINI.
JH..
P.
KlLL.Ahl.
WILLIAM
El Paw,
H.
M.D
51.1).
SC:RAG(;,
iU.1)
Teaas
Maternal unconjugated estriol levels ~LWW measured throughout the 28 to 41 week intenlal in two groups of accuratrly dated normal pregnancies. The first group consisted of randomly sampled pregnancies on which 285 unconjugated estriol determinations were prrformed. The logarithms of the mean ualues plotted into a positive sloping, relatively .straight line ulhich uus disrupted bl; a plateau orig-inating at 31 to 32 weeb and terminated at 35 weeks xlhrrc therr began a .steep surge to a point at 36 weeks (SUrgQ point) that returned value5 to fit the previously established .rtraight line. To Avestigate thesejindings in individual pregnancies, a second group nine subjects was studied with .serial unconjugated e.ytriol determinations. In all nine of these subjects, the Aurge point could be identified statistically and occurred at a mean gestational age C{ 35.0 + 0.6 (1 S.D.) weeks. Data,from the first group oJ mndomlp sampled pregnancies indica.te that the .curge point occurred around a mean gPvtationa1 aie of 36.0 weeks and was conJirmtd by data from the second group .srrially sampled individual subjects showirq th,r surge point as a statistically dejnable marker in normal pregnancies.
(rf
of
I N NORMAL pregnancy a continued increase in the rate of fetal adrenal growth occurs between 28 and 41 weeks’ gestation and becomes particularly marked between 34 and 36 weeks.’ This is associated with a sharp rise in circulating fetal and amniotic fluid cortisol levels.‘, ’ The exposure of fetal tissues to rising cortisol levels is probably the initiating event in a series of final maturation steps that include increasing pulmonary surfactant production and rising glycogen stores.“. ’ The mechanisms controlling fetal adrenal growth and steroidogenesis are largely modulated by the fetal hypothalamus and pituitary. ‘-” Accelerated fetal adrenal growth occurring during the 34 to 36 week interval has been linked to a concomitant rise in levels of circulating fetal pituitary prolartin.” In addition, rising cortisol levels are associated with a simultaneous
From the Deportment oj Olxtrtnc~ William Aeaumont Army Medical Rmived
Accepted
for
publicnlwn
December-
Suptembrl-
fall (suppression) in circulating fetal pituitary ACTH,‘O implying, as has been demonstrated in sheep, that the fetal adrenal (probably definitive cortex) has become increasingly sensitive to ACTH. ” Unconjugated estriol levels. as measured in the peripheral maternal circulation, are known to correlate closely with fetal adrenal steroidogenesis and the fetal corticotrophic mechanisms which regulate it. 7. ‘. I2 This consideration led us to study maternal unconjugated estriol levels in a series of uncomplicat.ed pregnancies at carefully documented intervals between 28 and 41 weeks to observe if known changes in fetal adrenal steroidogenesis would be reflected by alterations in maternal serum estriol concentrations.
Materials and methods Serum unconjugated estriol was measured in a series of normal, carefully dated pregnancies by a specific, rapid, nonchromatographic radioimmunoassay described previously.‘3 Serum was obtained by peripheral venipuncture and stored after separation at -20” C. until assayed. Each series of samples was run in the same assay such that between-sample variation was held to < 5.0 per cent. Blood samples were obtained between 13:30 and 15:30 hours from ambulatory outpatients with uncomplicated pregnancies delivering
arid Gywx~log~, Centw.
9. 1975.
16, 1975.
Reprint wqursts: .411rn P. Rillam, M.D., Department Obstetrics and Gynecology, William Beaumont Army Medical Center, El Pasn, TPXCIS 79920. Pwwnt addww: John Rcjproductive Biology, Canon St., Torm~r,
E. Buster, M.D., Diwsion Harbor General Hospital, Ccllifornin 90509.
oj 1000
oJ
K’.
672
Volume Number
Third-trimester
125 5
I
I
I
28
30
I 32
GESTATIONAL
34
)
)
I
,
36
38
40
42
AGE
serum unconjugated
estriol
673
(WEEKS)
Fig. 1. The means and standard errors for the means from 285 unconjugated estriol determinations between weeks 28 and 41 of gestational age on a semilogarithmic plot. Figures in parentheses indicate the numbers of samples for each point.
healthy term neonates. Pregnancies were dated on the basis of accurate and regular menstrual histories, confirmed by accurate midtrimester sizes, and reconfirmed by the pediatrician’s examination utilizing standardized published weight, physical, and neurologic criteria. i4-r6 Two groups of pregnant women were studied. The first group consisted of randomly selected subjects from whom one to nine samples were obtained between 28 and 41 weeks’ gestation, The great majority were sampled from one to three times for a total of 285 samples. The second group consisted of nine subjects who provided serial samples. Clinical data for these subjects are summarized in Table I. Data obtained from the first group of subjects were treated in the following manner. The means and standard error of the means from the 285 samples were placed on a logarithmic ordinate plotted against gestational age on a linear abscissa (Fig. 1). The logarithmic transformed mean values plotted into a positive sloping, nearly straight line disrupted by a plateau originating at the 31 to 32 week point and terminating at 35 weeks where a steep surge began and terminated at the 36 week point (surge point) that returned values to fit the previously established straight line. In order to confirm the above impression, a least-squares regression line with 95 per cent confidence limits was placed through all points between 28 and 40 weeks with deliberate exclusion of the 33 to 34 week and 35 week point (Fig. 2). The fit of this line was very close (r = 0.997). It was noted that the 33 to 34 week point (A) fell exactly on the 95 per cent confidence line and the 35 week(B) point well below it, indicating that probably neither point belonged to the straight line logarithmic transformed relationship described above. The 33 to 34 week point (A) and the 35 week point(B) were then compared by unpaired t tests with corresponding points predicted by the regression
Fig. 2. Logarithmic transformed regression line with 95 per cent confidence limits fitted to points between 28 and 40 weeks with deliberate exclusion of the points at 33 to 34 and 35 weeks where the numbers plateau from the trend of surrounding data points. The 33 to 34 week point (A), and the 35 week point (II), the 35 to 36 week interval (C), and the 41 week point (0) are tested statistically for both their relationship to the 95 per cent confidence intervals from the logarithmic transformed regression line and the corresponding values predicted from that line by unpaired t tests.
32282420 = 16. f 14p 12; IOB 6:, w 6-
$fSEY i--J-+ 28 30 32 14 36 38 40 42 GESTATIONAL
Fig. 3. Statistical regression line
AGE
(WEEKS)
definition of the surge point. A least-squares placed through the four points between 29.5
and 35 weeks and extrapolated
to week 36 point gives a
predicted future value ?95 per cent confidence first value falling outside of the 95 per cent interval is defined as the surge point.
limits. This confidence
line (Fig. 2). For the calculation, it was assumed that corresponding points had the same variance. The difference between points at the 33 to 34 week interval was not significant (p > 0.05) but the difference at 35 weeks was highly significant (p < 0.01). In order to investigate the significance of the surge point, the mean values and variances for the samples obtained to compute the 35 to 36 week points were compared by unpaired t tests, also confirming a highly significant difference (p > 0.01) (Fig. 2). The surge point was also defined another way (Fig. 3). A least-squares regression line was placed through the four points between 29.5 and 35 weeks and extended to week 36 with extrapolation of a future value at 36 weeks &95 per cent confidence limits of 11.8 -C 1.3 ng per milliliter. The measured level at 35 weeks of 15.3 ng. per milliliter fell
674
July 1, IY7ti Am. J. Obstet. Gynecol.
Buster et al.
L&o,
4imu 283032 343638 4042 :‘8 JL 24 1
GESTATIONAL
AGE (WEEKS)
Fig. 4. Individual plots troni nine pregnant sunjects with accurate dates. The surge point occurs III) earlier than 35.0 weeks and no later than 37.0 weeks with a inean of 86.0 2 0.6 (I SD.). In eight of nine subjects labor occurred between 39..i and 41.0 weeks with a tnean surge point to labor interval of 3.9 2 0.6 (1 SD.) weeks. In one subjec I (,J.I..), labor occurred at 3X.5 weeks with a surge point to labor interval of‘ 2.5 weeks.
well above the 95 per cent conhdence inter\-al predicted bv the line.” Data from the second group of subjects were treated in the following manner. Individual points were placed on a semilogarithmic plot exactly as described with the mean data from the first group. The surge point was then calculated and defined as follows (Figs. 3 and 4). With the use of the logarithms of the estriol concentrations from the first 3 weeks of observation a leastsquares regression line was calculated and plotted. The expected value and its upper 95 per cent confidence limit for the next week’s concentration were then calculated from the slope of the line. Whenever the actual observed serum concentration fell below this limit it was assumed to be a part of the previously calculated regression line and a new regression line was calculated incorporating this additional data point. The next actual concentration would then be compared to the predicted next upper 95 per cent confidence limit and the process was continued until the first measured concentration exceeded the upper 95 per cent conh-
dence limit. This point was considered as not part of this regression line and was defined as the surge point.
Results Data from the first group of pregnancies studied are shown in Fig. I. This figure depicts the logarithmic means and SXM. from 285 serum samples measured for unconjugated estriol levels plotted against the confirmed gestational age in weeks. The over-all positive logarithmic transformed linear slope and the transient downward displacement between 31 to 32 and 35 weeks occur as were noted previously. Although the point at 32 to 33 weeks is not significantly different (p > 0.05) from the corresponding value predicted by the regression line, this point falls on the lower 95 per cent confidence band for the entire line, implying that it probably does not belong to the straight logarithmic relationship described by the other points. The point at 33 to 34 weeks appears, therefore, to be transitional since it becomes significant if moved forward another O.:‘, weeks (p < 0.05).
Volume Number
Table
Third-trimester
125 5
I. Clinical
Subject J. C.
data from
nine pregnant
subjects
selected
for sequential
serum unconjugated
Gravida
PCWU
Fetal wt. at delivery (Gm.)
Gest. age at delivery (wk.)
27
4
1
2,948
41.0
9
Cord
40.5 39.5 39.5 40.0 40.0 40.0 40.0 38.5
9 10 9 9 10 9 10 9
None None None None None None None None
2.5
1
0
2,693
C. B. L. B. C. M. c. y. c. M’. G. R. J. L.
24 27 18 24 25 27 25
1 3 2 2 2 4 3
0 2 0 1 1
3,218 3,600 3,303 2,562 3,473 3,062 2,736
:
A second decline occurs at 41 weeks. This point falls below an extension of the 95 per cent confidence bands and is also significantly lower (p < 0.05) than the value predicted for 41 weeks by the regression line (Fig. 2). Fig. 3 depicts placement through the mean data points of‘ ;I least squares regression line as it is used to define rhe location of the surge point. Data f‘rom the group of nine individual pregnancies studied with serial samples is graphed on individual plots in Fig. 4 and tabulated in Table II. Fig. 4 depicts placements of regression lines, predicted values ?95 per cent confidence intervals, and the placement of the surge point as described. The mean gestational age at which the surge point occurred from all nine subjects was 36.0 rt 0.6 (I S.D.) weeks. The earliest placement was at 35.0 weeks and the latest was 37.0 weeks. In eight of nine subjects, spontaneous labor occurred between 39.0 ancl 40.5 weeks, and the mean surge point to labor interval was 3.9 -C 0.6 (1 SD.) weeks. In one of the nine subjects (J.L.), labor occurred at 38.5 weeks and was associated with a surge point to labor interval of 2.5 weeks.
Comment Numerous publications in discussing normal values for serum and urinary estriol during the third trimester have depicted a smooth curve which gradually swings upward until term.‘8-21 The 31 to 35 week plateau and 35 to 36 week surge described in this study have so far been unacknowledged. These phenomena may, in part, he unique to the unconjugated fraction of circulating estriol since this moiety is almost exclusively fetoplacental in origin and has not been shifted and metabolized through multiple maternal compartments as are the various estriol conjugates.“’ 23 Another explanation may lie with inaccurate dating of gestational ages in some series. Finally, in several publications the data has been condensed into 2 to 4 week intervals such that the rather short-term events described here would remain unnoticed.‘x-2’
APw
Complications
(5 min.)
compression during labor, otherwise none
Table II. Gestational age at time of surge surge point to labor interval Sd@
vi. C. L. C. C. c. G. J.
point
and
Gestational agg at surge point (wk.)
Surge point to labor interval (wk.)
37.0 37.0 35.5 35.0 35.5 36.0 36.0 36.0 36.0
4.0 3.0 3.5 4.0 4.5 3..5 4.0 4.5 2.5*
B. B. M. Q. w. R. L.
Mean ? 1 S.D.
675
studies
4-e
M. B.
estriol
36.0
f 0.6
3.9 t 0.6
*Labor at 38.5 weeks and not included in mean calculation. Any attempt to explain the downward deviations from an otherwise positive sloping logarithmic transformed line must be speculative with the present state of knowledge. It is plausible that these effects are due to increasing levels of fetal cortisol leading to decreased levels of ACTH.‘s ‘* “This in turn may cause decreased production of fetal dehydroepiandrosterone sulfate (DHEA-S) with a resultant decrease in placental estriol production. 7. 8 The 35 to 36 week estriol surge is more difficult to explain. The resumption of the previously established upswinging logarithmic transformed linear relationship may represent a sensitizing phenomenon such that the fetal cortex becomes more responsive to fetal ACTH and continues to produce increasing amounts of DHEA-S despite decreased circulating ACTH. Since circulating fetal prolactin rises sharply after week 34 and since it is believed to enhance fetal adrenal growth, lipid biosynthesis, and cholesterol (steroid precursor) storage, ‘it is possible that prolactin may promote increasing fetal adrenal zone sensitivity to ACTH through a permissive or additive effect. A similar situation may occur in the 40 to 41 week interval where the estriol curve slopes downward. The occurrence may reflect on further increases in fetal cortisol production and additional feedback ACTH sup-
676
Buster et al.
pression just prior to the onset of term labor.‘O In premature labor, an event associated with rising maternal estradiol levels and falling progesterone levels,” estrio1 concentrations have not been studied. However. the changes observed in estriol concentratiom at around 36 weeks may be more related to the steriod mediated maturation process active at that time rather than to the mechanisms triggering the onset of labor, as suggested by the findings in subject J. L.. who labored at 38.5 weeks, 2.5 weeks after the estriol surge poim. Determination of the surge point in individual pregnancies could be utilized as a noninvasive clinical test for dealing with patients \vho at-r first etKountet.ed during the third trirnesLe?- ~rntt drc corrrplicatrd b\ itiaccuraW or misleading riates. I‘he multitude of as well as frankI\ harartlous, probletns irritating, associated with “bad date” pregnancies are lvrll knowrt to all practicing obst,etricians. These data ittdicatc that. in il normal pregnatic v deVitr~d to deliver ,lt tcrnt. deterrninatior~ of tltrx surge poittt ~\ould permit: (1) an accurate c~stimation of’ the tcrrrt ~~;it‘tUriticJIl tlatc Lvithout other clinical irifortrla~iotl ;%II~ (2) intlicate thv
time at which adrenal steroidogenesis has ah.eaclv been shifted tol\xrd increasing corticoid produc tiorr and. 1~1 inference, increasitrg pulnionar\~ sttrfactatit produ(.tion. This latter point will cleat-l! require further, docurnentatioti. In order to utilize the surge poitil detet minatioti lorclinical purposes, it is essential that all sartiples be run through thr sa~ne assal in adjacent tubes so as to minitnizc rnrthodologir errors. It is, therefore. ot~t‘ \veekly blood samples f‘roln current prxtic-e to “bank” patients with poor dates with the expectation (4 running the entire series if. the need should arise. ‘The c;tlculation is best carried out with fout. or ntot-r points itt lvhich w plate a regression line. If three point> ;ttx‘ utilized. the regression line must fit WY\ closel\ in order to predic I ;I future value v;it tt ;I \vorkablv nat-roe !?5 per cent c,onfidence interval. Prelimittarv c-lirtical experience \\ith this approach has been cncx)uragirtg ‘I‘he authotx wish to thank 1)x.. Martin Vusynowity oi William Reautnont Army Medical Ctrnter ;uld Dr. John Rlarshall of Harbor General Hospital for their statistical assisraiicr ;iricl et~t‘o~irageiricttt.
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Data.
Philadel-
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