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Lead poisoning in monkeys during pregnancy and lactation
The Science o f the Total Environment, 30 (1983) 221--229
221
Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands
LEAD POISONING IN MONKEYS D U R I N G P R E G N A N C Y AND LACTATION*
P. TACHON, A. LASCHI, J.P. B R I F F A U X and G. BRAIN
Institut Pasteur Lyon Toxicologie, Domaine du Poirier, Lentilly, F 69210 L'Arbresle (France) P. CHAMBON
Institut Pasteur de Lyon, 77, rue Pasteur, F 69365 Lyon Cedex 2 (France) (Received October 2nd, 1982; accepted January 21st, 1983)
ABSTRACT Twenty-two female monkeys (Macaca irus) were given lead acetate by an intramuscular route during pregnancy and/or lactation. A high dose of lead (5 mgPb2+/kg/day) can induce abortions and death in pregnant monkeys. At a lower dose (1 mgPb2+/kg/day), the foetus can be exposed to lead via placenta and/or maternal milk. Studies using optical microscopy showed the typical renal lesions in mothers treated during pregnancy (five months) and an erythrodiapedesis in cerebral matter of the new-born at birth.
INTRODUCTION
The transfer of lead across the human placenta and its potential threat to the conceptus have been recognized for well over a century. In the past, w o m e n working in printing, and exposed to lead, showed a high percent of abortions, although t o d a y the incident rate is lower (Angle and McIntyre, 1964). Many workers consider that the human foetus is an organism that is especially sensitive to lead through maternal exposure; the placental transfer of lead begins as early as the 12th week of pregnancy (Cavalleri and Candura, 1979; Barltrop, 1969). Other studies have shown a significant correlation between blood lead of the new-born and of the mother (Alexander and Delves, 1981). Placental permeability to lead has been studied in conventional laboratory animals. At high doses by oral intake, lead induces a reduction in the number *Supported by grant 269-77-1 ENV F o f the Commission of the European Communities, Environmental Research Program. 0048-9697/83/$03.00
© 1983 Elsevier Science Publishers B.V.
222
of pregnancies in mice, and provokes an early mortality in mouse embryos before implantation (Jacquet, 1975; 1976; 1977; 1978). On the other hand, if the implantation does take place, the mice foetuses have a growth delay, an important post-implantation mortality and an increase of skeletal malformations (Jacquet, 1978; 1979). Also, lead induces a reduction of the blood flow in mouse e m b r y o s (Gerber et al., 1978). These experiments show that data on lower mammals often provide a good indication of the human response. But it is not possible, due to differences in placental structures and other factors, to extrapolate these data directly to human placental permeability (Carpenter, 1974). The use of primates permits an improvement in the prediction of the teratogenic hazard of many chemicals of pharmaceutical products (Bruggeman and Granwiller, 1976). Many workers have already identified several similarities between man and the m o n k e y , with regard to placental structure (Panigel et al. 1973), embryonic intra-uterine development (Hendrickx, 1971) and teratogenic sensitivity (Wilson, 1972). Therefore we have used primates in lead intoxication studies because of these similarities and because of the lack of clarity in extrapolating data obtained on lower animals to man. In order to study lead intoxication during pregnancy and lactation, we induced an experimental saturnism in female monkeys, and studied the passage o f lead from mother to foetus together with the histological changes arising from treatment with lead.
MATERIALS AND METHODS This study was carried out according to G.L.P. on 22 females Macaca irus in a genital activity period. Animals were kept in individual stainless steel cages and received a diet containing corn, oats, soya, bananas, skim-milk, sugar, fats, vitamins and beverages given ad libitum. Monkeys were caged for several months in order to steady the menstrual cycle. Gestation control
The biological gestation diagnosis was made in the mouse by injection of the urine of supposed fecunded females. This test was often confirmed by the post-implantation haemorrhage which appeared in our experimental conditions (Landsmann et al., 1978) after 17--30 days. Lead t r e a t m e n t
Pregnant m o n k e y s are dosed with lead acetate administered each day b y the intramuscular route. Animals are divided into several groups (see Table 1). The day preceding expected delivery, a caesarean was carried out for m o n k e y s of groups l a and 2a. For groups l b and 2b, after delivery,
223 TABLE 1 GROUPS OF MONKEYS AND THEIR LEAD DOSES (in mg/kg/day) Group
1
2
3
Monkey No.
Treatment period Gestation
la
35; 39
+
lb
23;26;41; 42;43
+
2a
27; 29; 31; 33; 40
+
2b
21; 30; 32
+
2c
28; 34; 38
3a
3;5;9;11
Dose
~+
mg Pb" ]kg/day Lactation
-b
-}+
+
new-borns were suckled by their mothers who continued to be treated. In group 2c, mothers were treated from delivery, and during lactation.
Lead analysis The lead dose was determined on mothers' blood and milk, and in foetal blood by atomic absorption spectrophotometry, for groups l a + b and 2a + b at birth or sacrifice day, and for group 2c from the first day of life and during lactation. The samples were diluted 1:4 with 0.1% solution of Triton X-100 in 0.1% nitric acid and injected into a graphite furnace (HGA 70) with atomization at 2100°C. For females that were sacrificed, bone lead concentration was examined in groups l a + b, 2a + b and 3a, by flame atomic absorption spectrophotometry, after mineralization of the samples.
Histopathological assays After the caesarean on m o n k e y s of groups l a and 2a, and at the time of the mother's death in group 2b, the mother's liver, kidneys, placenta and eight organs of the foetus or new-born (liver, kidneys, brain, eyes, skin, heart, adrenals, umbilical cord) were removed in order to undertake a histological examination. Organs were fixed in a formal solution (liver and brain) or in Bouin liquid (the other organs) for routine histopathologic processing and staining with Hemalum Phloxine Safran (HPS). In group 2c, at the end of experiments, a histological examination of the young m o n k e y ' s brain was carried out by a micro-optical technique. After pentobarbital anaesthesis, y o u n g m o n k e y s were bled and a perfusion was made, first with a physiological solution and then with glutaraldehyde products. The brain was then removed for routine histopathologic processing and staining with HPS (Table 2).
224 TABLE 2 HISTOLOGICAL EXAMINATIONS
Group
Monkey number
la
35 39
0 0
lb
23 26 41 42 43
270 150 150 480 480
27 29 31 33 40
0 0 0 0 0
2b
21 30 32
10 3 60
2c
28 38
150 150
2a
Age o f y o u n g s at sacrifice (days)
Histological e x a m i n a t i o n s
M o t h e r : liver; k i d n e y s ; p l a c e n t a F o e t u s : l i v e r ; k i d n e y s ; b r a i n ; eyes; skin; heart ; adrenals; umbilical cord
Young monkeys: brain
M o t h e r : liver; k i d n e y s ; p l a c e n t a F o e t u s / n e w - b o r n : liver; k i d n e y s ; b r a i n ; eyes; s k i n ; h e a r t ; a d r e n a l s ; u m b i l i c a l cord
Young monkeys : brain
RESULTS
Clinical observations For group 2a ÷ b the evolution of b o d y weight during pregnancy did n o t show any modification following the lead treatment. After three months treatment, the highest dose of lead (group 3a) was lethal for females during gestation. (Table 2). Foetuses were macroscopically normal: we did not observe any skeletal malformation. The gestation was also not affected by t r e a t m e n t at I mg Pb: +/kg/day. In this groups (2a + b), the mean weight of foetuses removed just before the delivery, or of the new-born, was ~ 300g (Table 3). After delivery, we continued to treat
TABLE 3 G E S T A T I O N D U R A T I O N A N D F O E T U S O R N E W - B O R N W E I G H T ( M e a n + SD) Group
Gestation duration (days)
F o e t u s o r n e w - b o r n w e i g h t (g)
l a ~- b 2a -b b 3a
1 5 0 . 9 +- 9.7 1 5 7 . 6 -+ 4.1 1 0 9 . 5 -+ 19.2
3 0 0 . 6 + 60.9 3 1 8 . 8 + 36.4 1 4 8 . 0 + 43.5
225 TABLE 4 LEAD CONCENTRATION ( M e a n + SD)
ON DAY OF SACRIFICE
OR AT BIRTH (NEW-BORN)
Group
Mother bone (mg/kg)
Mother b l o o d (pg/1)
Mother milk (p~g/1)
New-born b l o o d (pg/1)
la + b 2a + b 3a
21.7 + 19.8 98.4 + 67.9 2 4 0 . 5 -+ 1 8 4 . 6
5 0 . 5 -+ 23.4 1160.6 + 490.8
4 9 . 0 + 25.9 2220.0 + 622.7
4 7 . 0 + 26.8 1 3 8 6 . 7 + 510.1
mothers with the same dose of lead. In the first or second m o n t h after birth, mothers died, probably as a result of lead intoxication, and the newborn were sacrificed.
Lead dosages At the end of gestation, at sacrifice or at the deliver day the concentration of lead in mother's blood and milk and in the new-born's blood increased dramatically in the treated groups (2a + b) (Table 4). When females were sacrificed bone lead concentration in the groups 2a + b was high in comparison with the control group and reached 240 mg/kg in group 3a (Table 4). During lactation, for the treated group 2b, the blood lead concentration decreased in the new-born from 1837 + 55 pl/1 to 1038 + 270 pg/1. For treated group 2c, the lead concentration in mother's blood and mill~ increased rapidly and in mothers the milk lead concentration was always lower than the blood lead concentration. In young monkeys, blood lead increased slowly, but regularly (Table 5 and Fig. 1).
Histopathological examinations In treated mothers, after five months intoxication, we observed on the delivery day, or shortly afterwards, typical lead-poisoning lesions in the kidneys and intranuclear inclusions in the proximal tubular epithelium cells. No change was seen for placenta or liver, except for two females (No. 30 and 33) in which esinophil hepatocytes are noted. This eosinophilia was an impression of a start of cellular and tissular pain, the beginning of a non-normal state which could advance a necrosis. In foetuses and the new-borns, most organs were normal. In the brain, we noted an erythrodiapedesis in the cerebral m a t t e r before or just after birth. Young, five m o n t h s old (No. 28 and 38) monkeys, had a normal cerebral histology but with a mean blood lead concentration of 300 pg/1.
DISCUSSION
In the present study, we analyzed the influence of lead on the foetus through exposure to the m o t h e r m o n k e y .
226
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Fig. 1. Lead concentration in mother and new-born during lactation. We found that high doses (SmgPb2+/kg/day) of lead induced abortions and death in female monkeys treated during pregnancy. This observation is related with Angle's observation in women (Angle and McIntire, 1964), and also with earlier observations on the evidence of several abortions in w o m e n in lead industries (Cantarow and Trumper, 1944; Oliver, 1914). At a lower dose (1 mg Pb 2 +/kg/day), the treatment is well tolerated during pregnancy. We didn't find any teratogenic effect, which is in accordance with the WHO bulletin (WHO, 1978). A recent report of Miller on foetal rat development influenced by high maternal oral lead exposure, shows no teratogenic response or reduction in foetal brain DNA content, although significant amounts of lead crossed the placenta (Miller et al., 1982). We
228
demonstrated, however, that a foetus m o n k e y can be exposed to lead via placenta and maternal milk: the foetus or new-born's concentration of lead shows clearly some lead-poisoning, even if there is n o t any growth perturbation. With regard to histopathology, we found typical renal lesions in mothers treated for five months. Our findings are in agreement with results previously reported by Colle et al. (1980). These authors found distinct pathological changes in the proximal tubular epithelium cells with intranuclear inclusions in m o n k e y s treated for nine months. In another study, carried out under the same conditions as Colle's, we were unable to find (using electron microscopy) these lesions in monkeys treated for twelve months. However, tubules showed m a n y alterations, e.g. desquamation of cells and structural alterations (Tachon et al., 1981). Thus, we confirm the fact that intranuclear inclusions appear early (in less than one year), and disappear when the intoxication is longest ( E f t h y m i o u and Chaboux, 1975). This p h e n o m e n o n may result from a rapid death and substitution of tubular cells exposed to lead. On the other hand, we found an erythrodiapedesis in cerebral matter of the new-borns at birth, after the mother's t r e a t m e n t during pregnancy. But this alteration was not f o u n d after the mother's treatment during lactation. These observations must be related to Reyners' work in rats and monkeys (Reyners et al., 1980). In conclusion, lead crosses the placenta and is passed in milk. This p h e n o m e n o n may induce a very early intoxication of foetus during pregnancy, and then of the new-born during lactation. On the other hand, high doses of lead induce abortions, while lower doses induce an increase of blood lead level without teratogenic lesion. REFERENCES Alexander F.W. and H.T. Delves, 1981. Blood lead levels during pregnancy. Int. Arch. Occup. Environ. Health, 48: 35--39. Angle C.R. and M.S. McIntire, 1964. Lead poisoning during pregnancy. Am. J. Dis. Child., 108 : 436--439. Barltrop D., 1969. Transfer of lead to the human foetus. In: D. Barltrop and W.L. Barland (Eds.), Mineral Metabolism in Pediatrics. Davis, Philadelphia, pp. 135--151. Bruggemann S. and J. Granwiler, 1976. Le primate dans les essais de t~ratog~n~se. Sci. Tech. Anita. Lab., 1: 243--248. Cantarow A. and M. Trumper, 1944. Lead Poisoning. Williams and Wilkins Co, Baltimore. Carpenter S.J., 1974. Placental permeability of lead, Environ. Health Perspt., 7: 129--131. Cavalleri A. and F. Candura, 1979. L'esposizione professionale delle donne al piombo inorganico, Med. Lavoro, 5: 341--344. Colle A., J.A. Grimaud, M. Boucherat and Y. Manuel, 1980. Lead poisoning in monkeys: Functional and histopathological alterations of the kidneys. Toxicol., 18: 145--158. Ef t h y m i o u M.L. and C. Chabaux, 1975. Pollution par le plomb, influence sur le rein. In: Rein et Toxiques, XIV journees du groupement fran~ais des centres anti-poisons, Lyon 27--28 f~vrier, 85: 205--222. Gerber G., J. Maes and J. Deroo, 1978. Effect of dietary lead on placental blood flow and on foetal uptake of s-amino isobutyrate, Publication No. 1390 Euratom Biology Division.
229 Hendrickx A.G., 1971. Embryology o f the Baboon. Univ. of Chicago Press, Chicago. Jacquet P., 1975. Effects of death in mouse due to lead exposure. Experientia, 31: 1312--1313. Jacquet P., 1976. Effets du plomb administr~ durant la gestation ~ des souris C57B1. C. R. Seances Soc. Biol., 170: 1319--1322. Jacquet P., 1977. Early embryonic development in lead-intoxicated mice. Arch. Pathol. Lab. Med., 101:641--643. Jacquet P., 1978. Contribution ~ l'~tude de la t~ratog~n~se du plomb chez la souris. Thesis, Louvain, Belgium. Jacquet P., 1979. Effets du plomb sur les chromosomes et sur le d~veloppement pr~ et post natal. CEC report. Landsmann F., A. Collin, M. Laine and G. Mazue, 1978. Embryopathies induites par la thalidomide chez Macaca irus. Sci. Tech. Anita. Lab., 3: 233--242. Miller C.D., W.B. Buck, F.B. Hembrough and W.L. Cunningham, 1982. Fetal rat develo p m e n t as influenced by maternal lead exposure. Vet. Hum. Toxicol., 24: 163--166. Oliver T., 1914. Lead Poisoning, H.K. Lewis, London. Panigel M., A. Koren and R.L. Northrop, 1973. Experimental rubella virus infection of marmosettes. Lab. Anim. Sci., 23: 68--71. Reyners H., E.G. de Reyners, P. Tachon, A. Laschi and J.R. Maisin, 1980. Lead encephalopathy in the adult m o n k e y : An ultrastructural approach, Toxicol. Lett., 258 (special issue 1) 76. Tachon P. and A. Laschi, 1981. CEC report. WHO bulletin, 1978. Environmental Health Criteria. 3, Lead, WHO, Geneva. Wilson J.G., 1972. Abnormalities o f intrauterine development in non-human primates. Acta Endocrinol. Suppl., 166: 261--292.
221
Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands
LEAD POISONING IN MONKEYS D U R I N G P R E G N A N C Y AND LACTATION*
P. TACHON, A. LASCHI, J.P. B R I F F A U X and G. BRAIN
Institut Pasteur Lyon Toxicologie, Domaine du Poirier, Lentilly, F 69210 L'Arbresle (France) P. CHAMBON
Institut Pasteur de Lyon, 77, rue Pasteur, F 69365 Lyon Cedex 2 (France) (Received October 2nd, 1982; accepted January 21st, 1983)
ABSTRACT Twenty-two female monkeys (Macaca irus) were given lead acetate by an intramuscular route during pregnancy and/or lactation. A high dose of lead (5 mgPb2+/kg/day) can induce abortions and death in pregnant monkeys. At a lower dose (1 mgPb2+/kg/day), the foetus can be exposed to lead via placenta and/or maternal milk. Studies using optical microscopy showed the typical renal lesions in mothers treated during pregnancy (five months) and an erythrodiapedesis in cerebral matter of the new-born at birth.
INTRODUCTION
The transfer of lead across the human placenta and its potential threat to the conceptus have been recognized for well over a century. In the past, w o m e n working in printing, and exposed to lead, showed a high percent of abortions, although t o d a y the incident rate is lower (Angle and McIntyre, 1964). Many workers consider that the human foetus is an organism that is especially sensitive to lead through maternal exposure; the placental transfer of lead begins as early as the 12th week of pregnancy (Cavalleri and Candura, 1979; Barltrop, 1969). Other studies have shown a significant correlation between blood lead of the new-born and of the mother (Alexander and Delves, 1981). Placental permeability to lead has been studied in conventional laboratory animals. At high doses by oral intake, lead induces a reduction in the number *Supported by grant 269-77-1 ENV F o f the Commission of the European Communities, Environmental Research Program. 0048-9697/83/$03.00
© 1983 Elsevier Science Publishers B.V.
222
of pregnancies in mice, and provokes an early mortality in mouse embryos before implantation (Jacquet, 1975; 1976; 1977; 1978). On the other hand, if the implantation does take place, the mice foetuses have a growth delay, an important post-implantation mortality and an increase of skeletal malformations (Jacquet, 1978; 1979). Also, lead induces a reduction of the blood flow in mouse e m b r y o s (Gerber et al., 1978). These experiments show that data on lower mammals often provide a good indication of the human response. But it is not possible, due to differences in placental structures and other factors, to extrapolate these data directly to human placental permeability (Carpenter, 1974). The use of primates permits an improvement in the prediction of the teratogenic hazard of many chemicals of pharmaceutical products (Bruggeman and Granwiller, 1976). Many workers have already identified several similarities between man and the m o n k e y , with regard to placental structure (Panigel et al. 1973), embryonic intra-uterine development (Hendrickx, 1971) and teratogenic sensitivity (Wilson, 1972). Therefore we have used primates in lead intoxication studies because of these similarities and because of the lack of clarity in extrapolating data obtained on lower animals to man. In order to study lead intoxication during pregnancy and lactation, we induced an experimental saturnism in female monkeys, and studied the passage o f lead from mother to foetus together with the histological changes arising from treatment with lead.
MATERIALS AND METHODS This study was carried out according to G.L.P. on 22 females Macaca irus in a genital activity period. Animals were kept in individual stainless steel cages and received a diet containing corn, oats, soya, bananas, skim-milk, sugar, fats, vitamins and beverages given ad libitum. Monkeys were caged for several months in order to steady the menstrual cycle. Gestation control
The biological gestation diagnosis was made in the mouse by injection of the urine of supposed fecunded females. This test was often confirmed by the post-implantation haemorrhage which appeared in our experimental conditions (Landsmann et al., 1978) after 17--30 days. Lead t r e a t m e n t
Pregnant m o n k e y s are dosed with lead acetate administered each day b y the intramuscular route. Animals are divided into several groups (see Table 1). The day preceding expected delivery, a caesarean was carried out for m o n k e y s of groups l a and 2a. For groups l b and 2b, after delivery,
223 TABLE 1 GROUPS OF MONKEYS AND THEIR LEAD DOSES (in mg/kg/day) Group
1
2
3
Monkey No.
Treatment period Gestation
la
35; 39
+
lb
23;26;41; 42;43
+
2a
27; 29; 31; 33; 40
+
2b
21; 30; 32
+
2c
28; 34; 38
3a
3;5;9;11
Dose
~+
mg Pb" ]kg/day Lactation
-b
-}+
+
new-borns were suckled by their mothers who continued to be treated. In group 2c, mothers were treated from delivery, and during lactation.
Lead analysis The lead dose was determined on mothers' blood and milk, and in foetal blood by atomic absorption spectrophotometry, for groups l a + b and 2a + b at birth or sacrifice day, and for group 2c from the first day of life and during lactation. The samples were diluted 1:4 with 0.1% solution of Triton X-100 in 0.1% nitric acid and injected into a graphite furnace (HGA 70) with atomization at 2100°C. For females that were sacrificed, bone lead concentration was examined in groups l a + b, 2a + b and 3a, by flame atomic absorption spectrophotometry, after mineralization of the samples.
Histopathological assays After the caesarean on m o n k e y s of groups l a and 2a, and at the time of the mother's death in group 2b, the mother's liver, kidneys, placenta and eight organs of the foetus or new-born (liver, kidneys, brain, eyes, skin, heart, adrenals, umbilical cord) were removed in order to undertake a histological examination. Organs were fixed in a formal solution (liver and brain) or in Bouin liquid (the other organs) for routine histopathologic processing and staining with Hemalum Phloxine Safran (HPS). In group 2c, at the end of experiments, a histological examination of the young m o n k e y ' s brain was carried out by a micro-optical technique. After pentobarbital anaesthesis, y o u n g m o n k e y s were bled and a perfusion was made, first with a physiological solution and then with glutaraldehyde products. The brain was then removed for routine histopathologic processing and staining with HPS (Table 2).
224 TABLE 2 HISTOLOGICAL EXAMINATIONS
Group
Monkey number
la
35 39
0 0
lb
23 26 41 42 43
270 150 150 480 480
27 29 31 33 40
0 0 0 0 0
2b
21 30 32
10 3 60
2c
28 38
150 150
2a
Age o f y o u n g s at sacrifice (days)
Histological e x a m i n a t i o n s
M o t h e r : liver; k i d n e y s ; p l a c e n t a F o e t u s : l i v e r ; k i d n e y s ; b r a i n ; eyes; skin; heart ; adrenals; umbilical cord
Young monkeys: brain
M o t h e r : liver; k i d n e y s ; p l a c e n t a F o e t u s / n e w - b o r n : liver; k i d n e y s ; b r a i n ; eyes; s k i n ; h e a r t ; a d r e n a l s ; u m b i l i c a l cord
Young monkeys : brain
RESULTS
Clinical observations For group 2a ÷ b the evolution of b o d y weight during pregnancy did n o t show any modification following the lead treatment. After three months treatment, the highest dose of lead (group 3a) was lethal for females during gestation. (Table 2). Foetuses were macroscopically normal: we did not observe any skeletal malformation. The gestation was also not affected by t r e a t m e n t at I mg Pb: +/kg/day. In this groups (2a + b), the mean weight of foetuses removed just before the delivery, or of the new-born, was ~ 300g (Table 3). After delivery, we continued to treat
TABLE 3 G E S T A T I O N D U R A T I O N A N D F O E T U S O R N E W - B O R N W E I G H T ( M e a n + SD) Group
Gestation duration (days)
F o e t u s o r n e w - b o r n w e i g h t (g)
l a ~- b 2a -b b 3a
1 5 0 . 9 +- 9.7 1 5 7 . 6 -+ 4.1 1 0 9 . 5 -+ 19.2
3 0 0 . 6 + 60.9 3 1 8 . 8 + 36.4 1 4 8 . 0 + 43.5
225 TABLE 4 LEAD CONCENTRATION ( M e a n + SD)
ON DAY OF SACRIFICE
OR AT BIRTH (NEW-BORN)
Group
Mother bone (mg/kg)
Mother b l o o d (pg/1)
Mother milk (p~g/1)
New-born b l o o d (pg/1)
la + b 2a + b 3a
21.7 + 19.8 98.4 + 67.9 2 4 0 . 5 -+ 1 8 4 . 6
5 0 . 5 -+ 23.4 1160.6 + 490.8
4 9 . 0 + 25.9 2220.0 + 622.7
4 7 . 0 + 26.8 1 3 8 6 . 7 + 510.1
mothers with the same dose of lead. In the first or second m o n t h after birth, mothers died, probably as a result of lead intoxication, and the newborn were sacrificed.
Lead dosages At the end of gestation, at sacrifice or at the deliver day the concentration of lead in mother's blood and milk and in the new-born's blood increased dramatically in the treated groups (2a + b) (Table 4). When females were sacrificed bone lead concentration in the groups 2a + b was high in comparison with the control group and reached 240 mg/kg in group 3a (Table 4). During lactation, for the treated group 2b, the blood lead concentration decreased in the new-born from 1837 + 55 pl/1 to 1038 + 270 pg/1. For treated group 2c, the lead concentration in mother's blood and mill~ increased rapidly and in mothers the milk lead concentration was always lower than the blood lead concentration. In young monkeys, blood lead increased slowly, but regularly (Table 5 and Fig. 1).
Histopathological examinations In treated mothers, after five months intoxication, we observed on the delivery day, or shortly afterwards, typical lead-poisoning lesions in the kidneys and intranuclear inclusions in the proximal tubular epithelium cells. No change was seen for placenta or liver, except for two females (No. 30 and 33) in which esinophil hepatocytes are noted. This eosinophilia was an impression of a start of cellular and tissular pain, the beginning of a non-normal state which could advance a necrosis. In foetuses and the new-borns, most organs were normal. In the brain, we noted an erythrodiapedesis in the cerebral m a t t e r before or just after birth. Young, five m o n t h s old (No. 28 and 38) monkeys, had a normal cerebral histology but with a mean blood lead concentration of 300 pg/1.
DISCUSSION
In the present study, we analyzed the influence of lead on the foetus through exposure to the m o t h e r m o n k e y .
226
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150
ys
Fig. 1. Lead concentration in mother and new-born during lactation. We found that high doses (SmgPb2+/kg/day) of lead induced abortions and death in female monkeys treated during pregnancy. This observation is related with Angle's observation in women (Angle and McIntire, 1964), and also with earlier observations on the evidence of several abortions in w o m e n in lead industries (Cantarow and Trumper, 1944; Oliver, 1914). At a lower dose (1 mg Pb 2 +/kg/day), the treatment is well tolerated during pregnancy. We didn't find any teratogenic effect, which is in accordance with the WHO bulletin (WHO, 1978). A recent report of Miller on foetal rat development influenced by high maternal oral lead exposure, shows no teratogenic response or reduction in foetal brain DNA content, although significant amounts of lead crossed the placenta (Miller et al., 1982). We
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demonstrated, however, that a foetus m o n k e y can be exposed to lead via placenta and maternal milk: the foetus or new-born's concentration of lead shows clearly some lead-poisoning, even if there is n o t any growth perturbation. With regard to histopathology, we found typical renal lesions in mothers treated for five months. Our findings are in agreement with results previously reported by Colle et al. (1980). These authors found distinct pathological changes in the proximal tubular epithelium cells with intranuclear inclusions in m o n k e y s treated for nine months. In another study, carried out under the same conditions as Colle's, we were unable to find (using electron microscopy) these lesions in monkeys treated for twelve months. However, tubules showed m a n y alterations, e.g. desquamation of cells and structural alterations (Tachon et al., 1981). Thus, we confirm the fact that intranuclear inclusions appear early (in less than one year), and disappear when the intoxication is longest ( E f t h y m i o u and Chaboux, 1975). This p h e n o m e n o n may result from a rapid death and substitution of tubular cells exposed to lead. On the other hand, we found an erythrodiapedesis in cerebral matter of the new-borns at birth, after the mother's t r e a t m e n t during pregnancy. But this alteration was not f o u n d after the mother's treatment during lactation. These observations must be related to Reyners' work in rats and monkeys (Reyners et al., 1980). In conclusion, lead crosses the placenta and is passed in milk. This p h e n o m e n o n may induce a very early intoxication of foetus during pregnancy, and then of the new-born during lactation. On the other hand, high doses of lead induce abortions, while lower doses induce an increase of blood lead level without teratogenic lesion. REFERENCES Alexander F.W. and H.T. Delves, 1981. Blood lead levels during pregnancy. Int. Arch. Occup. Environ. Health, 48: 35--39. Angle C.R. and M.S. McIntire, 1964. Lead poisoning during pregnancy. Am. J. Dis. Child., 108 : 436--439. Barltrop D., 1969. Transfer of lead to the human foetus. In: D. Barltrop and W.L. Barland (Eds.), Mineral Metabolism in Pediatrics. Davis, Philadelphia, pp. 135--151. Bruggemann S. and J. Granwiler, 1976. Le primate dans les essais de t~ratog~n~se. Sci. Tech. Anita. Lab., 1: 243--248. Cantarow A. and M. Trumper, 1944. Lead Poisoning. Williams and Wilkins Co, Baltimore. Carpenter S.J., 1974. Placental permeability of lead, Environ. Health Perspt., 7: 129--131. Cavalleri A. and F. Candura, 1979. L'esposizione professionale delle donne al piombo inorganico, Med. Lavoro, 5: 341--344. Colle A., J.A. Grimaud, M. Boucherat and Y. Manuel, 1980. Lead poisoning in monkeys: Functional and histopathological alterations of the kidneys. Toxicol., 18: 145--158. Ef t h y m i o u M.L. and C. Chabaux, 1975. Pollution par le plomb, influence sur le rein. In: Rein et Toxiques, XIV journees du groupement fran~ais des centres anti-poisons, Lyon 27--28 f~vrier, 85: 205--222. Gerber G., J. Maes and J. Deroo, 1978. Effect of dietary lead on placental blood flow and on foetal uptake of s-amino isobutyrate, Publication No. 1390 Euratom Biology Division.
229 Hendrickx A.G., 1971. Embryology o f the Baboon. Univ. of Chicago Press, Chicago. Jacquet P., 1975. Effects of death in mouse due to lead exposure. Experientia, 31: 1312--1313. Jacquet P., 1976. Effets du plomb administr~ durant la gestation ~ des souris C57B1. C. R. Seances Soc. Biol., 170: 1319--1322. Jacquet P., 1977. Early embryonic development in lead-intoxicated mice. Arch. Pathol. Lab. Med., 101:641--643. Jacquet P., 1978. Contribution ~ l'~tude de la t~ratog~n~se du plomb chez la souris. Thesis, Louvain, Belgium. Jacquet P., 1979. Effets du plomb sur les chromosomes et sur le d~veloppement pr~ et post natal. CEC report. Landsmann F., A. Collin, M. Laine and G. Mazue, 1978. Embryopathies induites par la thalidomide chez Macaca irus. Sci. Tech. Anita. Lab., 3: 233--242. Miller C.D., W.B. Buck, F.B. Hembrough and W.L. Cunningham, 1982. Fetal rat develo p m e n t as influenced by maternal lead exposure. Vet. Hum. Toxicol., 24: 163--166. Oliver T., 1914. Lead Poisoning, H.K. Lewis, London. Panigel M., A. Koren and R.L. Northrop, 1973. Experimental rubella virus infection of marmosettes. Lab. Anim. Sci., 23: 68--71. Reyners H., E.G. de Reyners, P. Tachon, A. Laschi and J.R. Maisin, 1980. Lead encephalopathy in the adult m o n k e y : An ultrastructural approach, Toxicol. Lett., 258 (special issue 1) 76. Tachon P. and A. Laschi, 1981. CEC report. WHO bulletin, 1978. Environmental Health Criteria. 3, Lead, WHO, Geneva. Wilson J.G., 1972. Abnormalities o f intrauterine development in non-human primates. Acta Endocrinol. Suppl., 166: 261--292.