The human placental gradient for plasma insulin and blood glucose

The human placental gradient for plasma insulin and blood glucose WILLIAM

N.

FREDERICK BERYL

Z.

JOYCE

ELLS,

Minneapolis,

SPELLACY, C.

GOETZ,

GREENBERG,

M.D. M.D. M.D.

B.S.

Minnesota

T H E P LA C E N T A ’ s permeability t0 insulin has been investigated in previous studies with glucose levels, bioassays of insulin, and injections of radioactive insulin. These studies were either indirect, involved the use of a nonhuman insulin or were performed in experimental animals. The present study was undertaken to avoid these objections. The purpose of th is paper is to report levels of endogenous human insulin measured on each side of the placenta by an immunoassay technique. Blood glucose concentrations were simultaneously studied.

drawn from the antecubital vein of the mother and from the umbilical vein of the infant before the umbilical cord was clamped. The time interval between the drawing of the two blood samples was always less than 90 seconds. The blood samples were drawn without regard to the time of last feeding. The blood for glucose determination was placed in tubes containing sodium fluoride and potassium oxalate. The tubes were mixed well and immediately frozen at -20’ C. until analysis was performed. The glucose determinations were performed in duplicate by a method of Nelson and Somogyi.““~ 5o The blood for insulin determination was placed in tubes containing powdered heparin. The tubes were mixed well and immediately centrifuged at 2,000 r.p.m. The plasma was removed akd frozen at -20 C. Later both samples from one case were analyzed in duplicate in the same determination. The plasma insulin was assayed by radioimmunoassay method as described by Goetz.“‘, ”

Procedure Thirty patients were studied at the time of normal term delivery. All placentas were normal by gross and microscopic examination. Blood samples were simultaneously

Department of Obstetrics and Gynecology and Department of Internal Medicine, University of Minnesota Medical School.

Results

This study was supported by Grant No. Al556 from the National Institutes of Arthritis and Metabolic Diseases, United Statec. Public Health Service; from research funds of the Graduate School of the ITniversity of Minnesota; and by a grant from the Charles Pfizer aad Companies, Inc.

Blood glucose. The blood glucose values are shown in Table I and plotted in Fig. 1. It is seen from these values that there was a direct relationship between the blood glucose value of the mother and her infant across the placenta. The slope of the regression line for these points was 0.903 with a p value of 0.01. The mean umbilical vein blood glucose was 85 mg. per cent, and the mean maternal venous blood glucose was 109

Presented at the ]ohn L. McKeloej’ Twenty-five Year Anniversary Celebration, Minneapolis, Minnesota, Oct. 25, 1963; and at the Midwestern Section of the American Federation for Clinical kesearch in Chicago, Zllino6, Oct. 31, 1963.

753

754

Spellacy

Table I. 100 C.C. NO. 1 2 3 4 5 6 7 a 9 10 11 12 13 14 15 16 17 18 19 20 21 22 ‘3 2-l 25 26 27 28 29 30

et al.

Blood

glucose

Umbilical

values

ueinl

._

in mg. per

Maternal

vein

-

120 57

136 78

54 90 7-1 88 103 79 -

a9 115 10’ 109 119 111

54 72 78 92 133 96 98 62 74 68 78 127 73 136 80 48

a9 96 101 101 153 130 109 104 102 89 103 149 77 la0 102 72

mg. per cent. The individual infant’s umbilical vein glucose was always lower than the maternal value. Plasma insulin. The plasma insulin values are shown on Table II and are plotted on Fig. 2. It can be seen that the materna1 insulin values had a wide range of scatter. However, the umbilical vein insulin values were closely grouped in the 40 ,U unit per milliliter plasma range. There was no relation between the maternal and umbilical insulin levels. The slope for the regression line for these points was 0.005 with a p value of 0.001. The mean umbilical vein plasma insulin was 42 p units per milliliter, and the mean maternal insulin was 113 p units per milliliter. One dizygotic twin pregnancy studied had a maternal insulin of 245 p units per milliliter, with twin A level, 35 p units per milliliter, and twin B level, 21 p units per milliliter.

Table II.

Plasma insulin per cubic centimeter ....___.~~~ .-.__.~~..~-.--- NO. 1 L! 3

: i a 9 10 11 I:! 13 14

15 16 17 18 19 ‘0 21 “2 23 24 25 26 27 28 29 30

/ Llmbilical

values

uein 37 39 ,1-o 40 ‘II 1 28 :j 7 4” “7 “9 58 36 4” 3; 21 36 3-f 5:4 8-k 38 -13 4l! 66 36 50 23 56 47 ih 36

in ,U units

Maternal

rjein

72 45 85 145 115 a2 a9 14.5 45 166 186 56 91 35 ‘10 ‘NO 119 106 21 192 1’5 151 i-k 33 2iO 112 64 li4 a6 55

Insulin destruction studies with glutathione and cysteine were done. The umbilical vein insulin was completely destroyed thus supporting its identity as insulin.12 Comment

Blood glucose. Many experimental studies have been performed on a variety of animals, including human, to demonstrate the placental transfer of glucose. It was found that in rats, rabbits, and monkeys the fetal blood glucose is lower than the maternal glucose whereas in sheep and goats it is higher.4g 7f w 31 In human studies the umbilical cord blood glucose value is lower than the maternal value (umbilical cord blood glucose 94 per cent of maternal blood glucose in one study.z5. 54 Studies with C14-labeled sugars have demonstrated rapid placental passage.2’ After producing hyperglucosemia in the mother the fetal blood glucose rises

Volume Number

90 6

Plasma

Regression

line

slope

=

0.9035

insulin

Regresrion

and

blood

glucose

755

line slope = 0.0053

250

OI 50 Maternal

Fig. 1. Placental maternal pairs).

100 \/em

transfer

Blood

150

200

Glucose

mgm/lCQcc

of blood

glucose

250

I 50

I 100

Maternal

(cord-

but at a slower rate.35 In the present study it was found that there was a consistently lower blood glucose value in the umbilical vein blood but it was directly related to the maternal blood glucose. Plasma insulin. The problem of placental transfer of insulin was first studied indirectly. In the early literature it was postulated that insulin passed the placenta. The evidence for this was twofold. First, as clinical proof there was the association of infant pancreatic islet hypertrophy and the decreasing need for insulin during pregnancy in some patients with diabetes mellitus. This was supported in 1911 when Carlson and Drennan showed that pregnancy protected dogs from diabetes after pancreatectomy.5 Secondly, in several experiments in different animal species insulin was injected into either the mother or the fetus and the blood sugar was measured in both animals. A decreasing blood sugar in the animal on the opposite side of the placenta from the insulin injection was interpreted as showing placental transfer of insulin. InsuIin was thought to traverse the placenta of rats, goats, and sheep, but not that of rabbits.‘? ‘, “, 39, 40, a’~ ,13.4i, 45, js, 83 The objection of these experiments was that this is an indirect method of measuring insulin, and that by lowering

Fig.

2. Placental maternal pairs).

I 150

I 200

Vein Plasma Insulin

transfer

of plasma

,. I I I 250350 450

p units/cc

insulin

(cord-

the blood sugar of the animal injected one could also see a lower blood sugar in the attached animal because of the placental transfer of glucose without necessariIy the transfer of insulin. When radioactive-labeled insulin became available a more direct attack on this problem was begun. The first workers in this field were Goodner and Freinkel who used radioactive insulin and attempted to detect its passage across the rat placenta.“2 They found that rat placenta was relatively impermeable to insulin. Another study of rat placental transfer of P-insulin indicated that possibIy some transfer occurred.z8 It was found that I131-insulin would cross the rhesus monkey placenta but the umbilical cord concentration was only 20 per cent of the maternal IeveI. Later a study by Buse using radioactive insulin in humans showed only a minute quantity of the exogenous insulin passed the pIacenta1 barrier.3 SeveraI observers have found that the labeled insulin was “trapped” by the placenta and rapidly degraded, presumabIy by an enzyme “insulinase,“3>

17, 18, 23

With the use of a rat diaphragm glucose uptake bioassay method, Santos first measured insulin in maternal and cord plasma of 5 patients. 46 No insulin activity could be

756

Spellacy

et al.

determined in the umbilical cord plasma whereas the maternal plasma had a significant amount of insulin activity. It must be remembered that all of the previous studies were carried out in lower animals or used nonhuman insulin. The present study was undertaken to extend the results of Santos with a more sensitive and specific assay method. It is seen that insulin could be measured on both sides of the placenta but the maternal and -infant levels were independent of one another, thus tending to support a placental block to insulin transfer. Because the kinetics of insulin passage was not studied no statement can be made as to the dynamic exchange. Infants can produce insulin and newborn infants have been stimulated with intravenous glucose and have produced insulin as measured by the bioassay rat diathis insulin on phragm method. 1 Whether the fetal side of the placenta is produced by the fetus or transferred from the mother cannot be stated. One unlikely possibility is that the fetus destroys the transferred insulin as rapidly as it crosses the placenta. In effect the newborn was examined under varyina glucose concentrations and yet the measured insulin level was constant. This would indicate a relatively insensitive insulin-release mechanism to hyperglucosemia in the newborn. The area is being studied further. Clinically these facts have important significance for both the pediatrician and the obstetrician in reference to many previously held concepts. First, there is a constant fear of newborn hypoglycemia in infants of diabetic mothers.“, lo* 24 The hypothesis that this could be caused or potentiated by the dosage of maternal exogenous insulin received prior to delivery has been mentioned. Second, it has been postulated that exogenous insulin passage from mother to fetus could stimulate the production of insulin antibodies in the fetus.51 Third, recent experimental evidence has been produced to show that the normaI pregnancy is associated with hyperinsulinemia.32, 52 The source of this excess insulin is still unknown but one hypothesis is that the fetus could be supplying the excess insulin.

Fourth, the decreasing exogenous insulin requirement of approximately 10 per cent of the pregnant patients with diabetes mellitus has been postulated to be due to the transfer of a fetal insulin to the mother. This has been substantiated only by the finding of hypertrophied fetal islets of Langerhans.“‘, ” Finally, many extensive studies have found that babies born to mothers with diabet,as mellitus have an increased incidence of congenital anomalies.“” In animal studies it has also been shown that insulin is teratogenic.“’ 29 It has been postulated that, perhaps. the mother’s exogenous insulin passes the placental barrier and produces these fetal anomalies. All of these concepts will havtx to be re-evaluated in view of this and olhel experimental data supporting a placental block to the passage of insulin. The next obvious question is why does not insulin pass the placental barrier? This barrier will allow most substances with a molecular weight of 500 to 1,000 to pass freely by diffusion.l”s Ri Its permeability is not governed solelv by diffusion as stereoisomers of simple sugars have different rates of pasmolecular weight substances sag-e.I’ Hither ,_ such as antibodies in the S-J fraction will also pass.8 The S-i gamma globulins ha\-r: a molecular weight of 160,000.4’ Besides molecular size the permeability is governed by the molecular chargr, and for some substances by an acti\,e transport mechanism.“, “3 ‘I’. ” Insulin has a minimum molecular weight of 6,000 and an isoelectric point of .5.3.16. iii At a pH of 7.4 the molecule possesses a net negative= electric charge. It is now known that the insulin molecule ma\ form polymers with molecular weights of 48,000 or more.‘” The large size and molecular charge of insulin polymers could impede their placental passage. Thus, four possible explanations for the measured insulin difference would include: i 1) placental block : i 2) rapid placental destruction ; (3) placental binding: or (4) protein-insulin binding.

Conclusion Plasma insulin and blood been simultaneously measured

glucose have in maternal

Volume Number

90 6

Plasma

venous blood and umbilical vein blood at the time of a normal term delivery. Blood glucase appears to pass between the mother and fetus. Plasma insulin is present on both sides of the placenta but the concentration is lower on the fetal side and is unrelated to the maternal level. These results are inter-

insulin

and

blood

glucose

757

preted as evidence that natural human insulin, as measured by the immunoassay technique, does not freely pass the placental barrier. We are indebted to Mrs. G. Rapley and Mrs. M. Condon for their technical assistance.

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