Prolactin in normal pregnancy: Relationship of maternal, fetal, and amniotic fluid levels

Prolactin in normal pregnancy: Relationship of maternal, fetal, and amniotic fluid levels J. G S C H E:\1 K E R. \1. D. )\(

BF~·DA\"ID.

PH.D.

\V . Z . P 0 Ll S H L K . !\I . D . jl'nllllll'lll.

hmri

SrTI/111 prolactin ref/.\ 11/t'a.,mnl 'imulfi!II!'IJIISIJ b)' 11 homolugou., mdiuimmutwa.\.1/IY in I.J2 parlurimts and lhf'ir ofj.ljJring at tam. and in maltrnal serum and mnniot/,fluid during <•arious stag·,·s of normal jlngiWIII)'· Strum ltl'l'l\ oj 2()H ::t: 8 11g. fltr mil/ilitn m·rc flmnd in matt•nwl blood: tht cul'lf'.ljlmtdmg hT!'/.1 in th,• {'(J/'d S!TIIm
ihf' torrnj)f)llding !tr•tls of lhl' uwthn:'. Tlu• jJoS.\Ihlt .11111/'tl' of 1111' 1111111/11/ltjfuid jJm!arflll molau/,•, "hith 1s 1(/rntiml with ihl' human pituil11rr molrrulr·. i,, dilti/.1.\t'tl.

REc~.~ 1

S'l l'OIES

culminated in the isolatio11 ol

Material

separare growth hormone and prolactin in man.'· 2 The

Prolactin lcvds \\'ere determined in the following ;,ourccs: ( l J serum from 12 normal cvding \\Olllen and

abilit\ to quantitate these hormones b\ sensitin· hiologic'1· 1 and radioimmunologic assay techniques"· B have Jed to a definition of their activit\ in a \'ariety of

phvsiologic and pathologi< wnditions. 7 •

!l men: (:!J I H·t blood serum samples obtained from 92

panurients with an ti!Homplicated antenatal course and from their offsprin).i: 1he \\omen had ddinTed -;pomaneoush at term and there 1\·<·n· -IS male and +4 female ofbpri11g: (:\) maternal serum and anmioti<

The finding ol circulating prola;ludied tlw "

fluid prolanin determined simulraneoush in :.!9 hcaltll\ pregnant wona·n in different stages of gesta-

maternal and newborn serum prolactin kn·ls at the time of dcliven and the levels of prolactin in thl' amniotic fluid and serum during· difter!'nt stage~ of normal pregnann. It was fr>Lmd I hat pituitarv la
tion.

Methods

Preparation of samples. A ::, to I 0 m!. sample of

aninttcd throughout pregnann''' and serum le1-cls

maternal hlood was obwined at cleliven from th<· antecubital n·in. Blood from newborn infants was "it hdr;n\n from the umbilical wrd immediareh after deli1en. Amniotic lluid Kas obtained b' anmim<·m<·sis except in two cases. 1!1 one case ol H IH-cks' pregnann it I\ a~

during dilferent stages of gestation and puerperium 1\'l.'lT reported. 10 The prolactin molecule. identical with that f(ltllld in piruitan and serum. \\·as detected and reccntlv isolated from amniotic lluid. 11 • 12 The source of the larg-e amounts of amniotic lluid prolactin is ;;lill not dear. The correlation bct\IC·cn prolactin !nels i11 maternal and cord serum and those in the amniotic Huid i> as n·t nor knrmn.

obtained lrotn a 11\stcrcctonn specimen. In auotlwr case. at II weeks' gestation. th(' amniotic sac wa~ punctu1·ed l(ll· >ampling at the time of spontaneous

;tbortiotl. The blood "as allowed to dot and the seru nt ''as separated at ·F C. b\ centrifugation ( 1.000 g-. l.i minute~). Serum and amniotic fluid \H'IT stored at --:!00 C until ass;n·ed bv a homologous 1 adioimmunoas~

Fmm thr Depw1mn1ts of Obstetnr1~Gynerulugy and Pharmarololfl, HPbrew Unil•n~1it.v. Hadas.mh JVf,•diwl Ctnlt'r.

Rnri<•NI fur pahlimtion S!'ptnnhl'l 2·1, /'!/ .J

'ia\. :\ I 00 j-<1 quanti11 of I: I 0 dilution (I per celll ho\ille ~num albumin-pho,phmaline huller [ BSAPSB J 11 a;, etnplon·d in each tube as-a1. Radioimmunoassay. Human prolactin c•nnposcd r,f

At'ajlltd Dn·tmher 23. J97.J. Rr•{mnl rl'que
Jnumlnn, lsrad.

834

\'olume 12:1 '
Prolactin in normal pregnancy

o

•

835

Cord serum Maternal serum

) N° of Pairs

700

(3)

(15)

(15)

(21)

(16)

( 6)

( 16)

l

I 500 c

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0 ..... 0 '-

0..

300

100

/

l 0

5'o

50I 100

100 I 150

150 I 200

200 I 250

250 I 300

300 I 1000

Mother's basal serum Prolactin levels Fig. 1. Correlation of maternal and cord serum prolactin

isohormones B and C. isolated from amniotic Huid, 12 was used for radiodination and also served as a reference hormone. Rabbit anti-human prolactin was kindly supplied by the 1\IAMDD. Bethesda. Man·Jand. Coat anti-rabbit immunoglobin of Miles-Yecla, Rehovot, IsraeL served as a second antibody for the double-antibodv RIA. Iodination \\·as carried out with rz:q· by the method of Greenwood and associates 1a as follows: To l rn( ~i of 125T were successively added: 25pJ of 0.5M phosphate buffer (pH 7.6): 2fl,g of hPRl in 2:jft] of (l.O I M N ll~HC0;3 solution (pH S.l ): and I Of~-l of a (5 mg. per milliliter) chloramin-T solution freshlv preparn! in phosphosaline buffer (0.1 M, pH 7 .6). Instant and continuous shaking of the reaction tube followed for 40 seconds, until the reaction was stopped bv addition of ~!:"ip.l of sodium metabisulfite solution in PSB ( 12.5 mg. per milliliter). Separation of the labeled intact protein from damaged molecules and free iodine W
le~els

at term delivery in normal gestation.

ranged from HO to \[>0 mC:i per milligram. The first incubation (included 0.1 mi. of BSA-PBS. 0. I mi. of reference hormone or diluted serum, and O.l.'i mi. and antibody I :200,000 final dilution in 4 per cent 1\'RS in PSB) was performed for 24 hours. Then 0.15 mi. of labeled hormone. 8,000 to 10,000 c.p.m. in I per cent BSA-PSB, was added, followed by incubation t(n· I!) hours at 3 7o C. Then 50 Ml of goat anti-rabbit gamma globulin in 200 p.l of PSB was added and left at room temperature for 1.5 to 2 hours to allow the antibodybound labeled hPRL to precipitate maximally. All tubes were centrifuged at 2,500 g for 20 minutes at 25° C. and the supernatant \\·as discarded. Precipitate was counted in a Packard Autogamma Spectrometer. The lowest concentration of prolactin which could be accurately determined was 0.5 ng. of reference prolactin per tube. All assays were performed in duplicate. Log doses of reference hormone wne plotted against the logit. of B/Bo x 100 (bound wunts of the sample divided by bound counts when no cold hormone was added) in order to obtain a straight line rune. The method was found to be specific to hPRL. Neither human placental lactogen (HPL) nor human pituitary growth hormone (hGH) was detectable in the assay

I h-< t·ntlwr I:). ! q7_)

.-c-erMer. :'en-iJavic·, and Polishuk

.\111.

I

900~

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c:

!

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~Amntofic flutd

I

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~

400

300~

• • • • • •• • • A. • • • al 20 25 5

100

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15

A.

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•

...

1

• •

• .... e •• • • ~ • • • I •• •

ttl

a..

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...

0

0

Ill.

...

700~

en

... -

......

.A

- 600t

Oh~tct. (;\'lit>!

* ....

t

E

.1-

I

30

1

35

~ 40

Weeks of Pregnancy

Fig. 2. Prolactin len· Is in maternal '<'rum and in amniotic fluid during Yarious stages of pregnann.

system. Moreover, large doses of human folliclt:>-stimulating hormone (hFSH) and human tbvroid stimulating hormone (bTSH) ,,·ere complete!\ ineffective in inhibiting the reaction of labeled hPRL \\ ith antihPRL.

Results Serum prolactinlnels ranging from-! to :W ng. 1\'tTe found in normal adult males and in normal non pregnant IUJ!Ilen throughout the crcle. In uncornplicatccl pregnann at term. the prolactin len·! in maternal serum was 20M ± K S.E. ng. pcT miiiiiiter whereas ineis of ;Fi-! ± 14 ng_ per milliliter were found in the cord ~erunl. Prolactin SLTurn le\·el~ in each of a cord and maternal samples are jointh· depicted in Fig_ I. The cases were di\'idnl into 1arious groups according to the

mother's basal serum prolactin le\'ds. In 79 per cent of all prolactin estimations of maternal and cord serum at deliH·t-v. the cord 'erum prolactin len~ls were higher than the mother's. In the group in 11 hidt the mother's basal snum prolactin In ds 11 t·re helo\1' ::250 ng. per milliliter cord serum prolactin was higher in KK per cent of the pairs studied. In the remaining I :Z per cent. serum coni prolactin le\'els were similar to-or not significantlv lower than-mother's serum pn1lactin. \!\'hen the basal serum prolactin level of the mother exceeded ::200 ng_ per milliliter. in eight out of 19 pairs. coni serum proiactin were signihcantiv iower than that of the mother. ~ o uHTclation ,,-as f
\'olurne I ~:l

Prolactin in normal pregnancy

837

~umber~

placenta. Sex of the neonate showed no influence on the distribution of prolactin levels in corresponding sera. Fig. 2 shows prolactin concentrations in the amniotic fluid and maternal serum during different stages of normal pregnancy. There is a rise of prolactin serum levels of the mother with advancemem of pregnancy, whereas there seems to be a relative decline in the amniotic fluid prolactin tovvard the end of pregnancy. In 28 of 29 cases studied. the concentration ot amniotic fluid prolactin was 2- to 10-f()ld higher than that oft he mot her. In only one case of earlv pregnancy (8 weeks) was amniotic fluid level lower than corresponding mother serum level.

Comment Our finding of a progressive rise in prolactin levels during pregnancy. reaching peak levels near term, is similar to those previously reported. 11 This increase may be due to the increase in number and activity of lactotropes in the pituitary of pregnant women. 9 · 14 The role of increased prolactin in pregnancy is still not clear. Its rise parallels that of human placental lactogen. which may indicate that the principal fumtion of both is lactogenic. although other functions mav be assigned fi>r each. The problem of whether or not maternal prolactin is the sounT for the amniotic fluid prolactin is not resolved. From at heoretical point of view the source of the huge amounts of prolactin in amniotic fluid might be: (I) the fetal pituitary gland, (2) the placenta. or (3) the mother's pituitary. The fact that considerable amounts of prolactin were found during the first trirnesler of pregnant) in the antniotic fluid excludes the first possibility. since at this stage the fetal pituitary secretes no or \en little prolactin. In the human subject, no data are available to demonstrate whether the accumulation of prolactin in the amniotic fluid is brought about via a selective passage of the hormone, originating in either the n1other or the placenta~ Josimm·ich and associates 15 assumed that in the rhesus

monkey the ongm of amniotic fluid prolactin is the maternal circulation. This accumulation is not a common phenomenon in all mammals. In the rat. f(>r example, considerable amounts of prolactin an· found in the maternal circulation but not in the amniotic fluid, 16 despite the fact that small amounts of iodinated prolactin are capable of passing from maternal cirndation to amniotic fluid and vice \ersa. 16 Results of the present study den1onstrated that in human subjects prolactin was present in amniotic Huid in all cases studied throughout pregnancv except of one case of early pregnann (8 weeks). Its leH·Is \\ere higher by 2- to I 0-fold than the corresponding maternal serum levels. Moreover, at term, it \\'as found that in ill per cent of cord prolactin levels were higher than the corresponding maternal serum. Since identical prolactin Illl>lecules were found in amniotic Huid, serum. and pituitary. 1 ' it is likely that the source of AF prolactin is the maternal circulation. If this assumption holds true, then one must assume that there exists a different rate of prolactin transfer from the mother via fi:tal ( ompartments to amniotic sac from one side and the opposite way from the other. The physiologic significance for the increased level of prolactin in the fettrs neonate, and amniotic fluid arc still not dear. It seems, from experimental studics, 17 that prolactin is crucial during fetal development. Data obtained from se\cral studies performed in human beings and in laboratory animals 1M· 1 ~ indicate that prolactin is involved in the regulation of sodium and waier. Also, prolanin was found io alter the permeability of gill membrane so that salt fish can adjust to fresh water '"·hen they inigrate for spa,,·ning. Hence it is possible that in the human subject during leta! development prolactin is essential f(n· osmotic regulation. We are greatly obliged to Dr. R. W. Bates. N IAMDD. for the hPRL antibody supply, and the devoted technical assistance of Michal Taasan is appreciated.

REFERENCES I. Hwang, P., Guvda, H .. and Friesen, H.: J. Bioi. Chem. 247: 1955, 1972. 2. Lewis, U . .J.. Singh. N. D., and Seavy, B. K.: Bioch. Biophys. Res. Commun. 44: 1169, 1971 3. Loewenstein . .J. E., Mariz, I. K., Peake, G. T., and Daughsday, \'\'. H.: J. Clin. EndocrinoL 29: 1383. 1969. 4. Franz, A. G., Illeinberg, D. L., and Noel, G. L.: Recelll Prog. Horm. Res. 28: 527, 1972. 5. Hwang, P., Guyda. H., and Friesen. H.: Proc. Natl. Acad. Sci. 68: 1902. 1971. 6. Jambs. L. S .. Mariz, I. K., and Daughaday, W. H.:]. Clin. Endocrinol. 34: 4H4, 1972.

7. Yuen, B. H., Keye, W. R .. and .Jalfe, R. B.: Obstet. GvneG>i. Surv. 28: 527. 1973. 8. D~whurst, C. S.: In Year Book of Ob>tetrics and Gynecology, Chicago, 1973, Year Book Medical Publishers, Inc.. p. 133. n

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L- •• ..-L.r..-:..-..--1

J. .Jill\ I\_ I Ill' JJ,

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1969. 10. Tyson,]. E.. Hwang. P .. Guyda, H., and Friesen, H. G.: AM. J. 0BSTET. GYNECOL. ii3: 14, 1972. I I. Ben-David, M.. Rodbard. D., Bates. R. 'vV .. Bridson, W. E .. and Chrambach, A.: .J. Clin. Endocrinol. Metab. 36: 951. 1973.

838

Schenker. Ben-David, and Polishuk

lktcnlhcJ L+). :\JJL

i :.'. Ben-Da\ id. '\I., and Chrambach. J\.: Endocrine Res. Commun. 1: 1\!'\, 1974. l.'l. Greenwood. F. ( :.. Hunter. \\'. \1.. and (;Jmer, .J. S.:

Riochem . .J. 89: 114. 1963. II. l'astct'ls . .J. L., Gaussett. A .. Dani{U). A .. Fctors. F .. :--Jicoll. C. S .. and \'aradllldi. 1'.: .J. Clin. Endocrinol. \Ictab. 34: 95'1.1!172. 1:-> . .Josimo\ich . .J. B .. \Yei;, (;., and Hutchison. ll. L: Endcwrinolno-'1ll7-l . ..... Q.::t~ 1. ';\f;4 .. . ...... I ti. Bcn-DaYid, 1\1., Schenker . .J. (;., Hammt'!. L. and ~

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l'oltshuk. i\'. Z.: Prcscntt'd at the international Snnposiurn on Growth Hormone and Related Peptides. Milan, ltah. September. 1975. 17. Sinha, Y. !\ .. Lewis. l' . .J .. and \'andnla11. W. P.. /11 Filtv-Third \lceting of the Endocrine Society, San Fra;His:!. 1!1/1.