International Archives of
Int Arch Occup Environ Hlth 40, 33-36 (1977)
Culutlional aNul Eriumnental (k
Health
©)Springer-Verlag
1977
Mobilization of Lead During Pregnancy in Rats* J.P Buchet, R Lauwerys, H Roels and G Hubermont Unite de Toxicologie Industrielle et Mdicale, University of Louvain, 30 Clos Chapelle-aux-Champs, B-1200 Brussels, Belgium
Summary Lead was administered to three groups of 4 weeks-old female rats at the dose of 1 ppm in drinking water The first group received lead continuously for 150 days before mating (with non-lead-treated male rats), during pregnancy and during 3 weeks after delivery In the second group lead was also administered for 150 days before mating but was withdrawn during pregnancy and during the post natal period In the third group lead was administered for 150 days; the animals were then kept without lead exposure during the 50 days before making them pregnant, during pregnancy and during the post natal period A control group received only demineralized water. On day 21 after delivery the mothers and their newborns were sacrificed and various parameters of blood-lead concentration (Pb-B), hematocrit (Htc), hemoglobin (Hb), free erythrocyte porphyrins concentration (FEP), 6-aminolevulinate dehydratase activity (ALAD) and tissues ALAD, free tissue porphyrins (FTP), lead concentration (Pb-T) were determined. In mothers and in newborns lead concentration in blood and in soft tissues were increased over control values only in the group in which lead had been stopped for 50 days before pregnancy This lead accumulation in soft tissues was associated with an increased FTP in liver, heart and brain of the mothers and in liver of the newborns This observation suggests that lead stored in the organism can be mobilized during pregnancy. Key words: Lead in blood Lead in tissues ALAD in blood ALAD in tissues Porphyrins in blood Porphyrins in tissues Transplacental transfer We have previously studied the effect of short-term oral administration of lead on various parameters of the heme biosynthetic pathway in pregnant rats and their offspring (Hubermont et al 1976) Three doses of lead ( 10, 1 and 0 1 ppm in drinking water) were administered for 3 weeks before mating, during pregnancy and during * Supportedbyagrantfrom the Commission of the European Communities (Project no 035-74-1Env B) and the Fonds de la Recherche Scientifique M 6dicale (Belgium)
34
J P Buchet et al.
three weeks after delivery No significant biological changes (except increased lead mothers concentration in blood and in kidney in the 10 ppm group) were found in the (ALAD) dehydratase -aminolevulinate 6 group ppm 10 but in the newborns of the porphyrins activity in blood and in kidney was significantly decreased and free tissue values. control over increased significantly was kidney (FTP) concentration of We thus concluded that the developing organism is more susceptible to the biothe logical action of lead than is the organism of adult animals and on the basis of preduring administered lead of level no-effect the biochemical parameters studied whether tested now have We ppm 1 around is gnancy and in the neonatal period 1 ( ppm) to female rats before longterm (150 and 200 days) administration of lead level found during the short-term ,no-effect" the making them pregnant will modify of the animal could be mobilized life early during study and whether lead administered pregnancy. subsequent a and transferred to the foetus during Four weeks-old female Sprague-Dawley rats were used Lead was administered as Pb(NO 3 ) 2 at the dose of 1 ppm in demineralized drinking water There were 4 groups of 8 female rats: the control group (a) received only demineralized water, 4 animals during 150 days and 4 during 200 days; since no significant difference in the biological measurements was found between the two subgroups, the results were pooled for purpose of comparison with groups (b), (c) and (d); group (b) received lead continuously for 150 days before mating (with non-lead-treated male rats), during for pregnancy and during 3 weeks after delivery; in group (c) lead was administered post the during and pregnancy during withdrawn 150 days before mating and was then natal period; in group (d) lead was administered for 150 days; the animals were during pregnant, them making before days 50 the kept without lead exposure during pregnancy and during the post natal period At time of sacrifice the animals in this group were thus 50 days older than those of groups (b) and (c). Before mating the Pb-B levels of the animals in group (d) were determined. Not all the animals became pregnant and in order to have a similar number of animals in each group only 4 pregnant animals were examined in each group At delivery, only 8 pups per mother were kept alive On day 21 after delivery the animals were homosacrificed Blood collections, whole body perfusion and preparation of tissue tissues and blood The 1976) al et (Buchet previously genates have been described of 3 pups were pooled to obtain sufficient sample volume for all the determinations. beFor each mother two ,,newborn" values were thus obtained which were averaged whole on performed were analyses biochemical following The fore statistical analysis blood: ALAD activity (in nmoles -ALA transformed/min x ml erythrocytes), hematoml crit (expressed in %), free erythrocyte porphyrin (FEP) concentration (in g/ 100 conand erythrocytes), hemoglobin (Hb) concentration (in g/100 ml whole blood) On the centration of lead in blood (Pb-B) (expressed in gg lead/100 ml whole blood) measurements whole homogenate of liver, kidney, heart and brain, the following mg protein), were performed: ALAD activity (in nmoles 6-ALA transformed/min x (expressed (Pb-T) tissue FTP concentration (in ng/mg protein), concentration of lead in methods analytical in ng lead/mg protein) and concentration of tissue protein The used have been described previously (Buchet et al , 1976). older The treatments and the slight age difference between the groups (group d mothers the of weight average than groups b and c) had no significant effect on the
35
Mobilization of Lead During Pregnancy in Rats
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at sacrifice (individual values ranging from 290 to 320 g) and of their pups (individual values ranging from 38 to 48 g). In mothers lead concentration in blood and in tissues was lower in the groups receiving lead continuously (b) or up to the beginning of pregnancy (c) than in the group (d) in which lead had been stopped for 50 days before pregnancy (Table 1).
36
J P Buchet et al.
In this group lead concentration in blood and kidney was significantly increased over control value This observation suggests that lead stored in the organism can be mobilized during pregnancy. This hypothesis is reinforced by the fact that just before mating the Pb-B of the female rats of group (d) were in the normal range (< 3 g/ 100 ml whole blood). In the same maternal group (d) FTP in liver, heart and brain were significantly increased (Table 1) but ALAD was not inhibited in blood or in tissue There was no effect of lead treatment on Htc, Hb and FEP level. In newborns, the change in lead concentration of blood and tissues paralleled that found in mothers (Tab 1) A significant increase was found in blood, liver, kidney, heart and brain of newborns of group (d), i e , that in which Pb-administration to the mothers was stopped 50 days before pregnancy (One way analysis of variance p < 0 05). The increased lead levels found in group (d) were not associated with a reduction of ALAD in blood or tissues but FTP of liver were significantly increased (p < 0 05, Table 1) Like in mother, no change in Htc, Hb and FEP level were found. In summary, the most significant observation made during this preliminary investigation is the increased accumulation of lead in newborns of mothers which had received 1 ppm Pb for 150 days and had been on a lead free regime for 50 days before pregnancy (group d) This lead accumulation was associated with an increased FTP concentration in liver of newborns It is surprising that this phenomenon was not observed when 1 ppm lead was administered continuously up to the beginning of gestation (group b) or throughout pregnancy (group c) since the lead body burden of these animals was certainly higher than that of group (d) This could be due to a reduction of the osteocytic or osteoclastic resorption of bone in both groups (b) and (c) (Eisenstein and Kawanoue, 1975 ; Hsu et al 1973) Lead inclusions have been described in osteoclast (Hsu et al , 1973 ; Von Mullem and Stadhouders , 1974) This could be accompanied by a reduction of their metabolic activity In group (d), the reduction of lead concentration in bone during the 50 day-period without lead administration before mating might have been sufficient to restore the osteoclastic activity. Further informations are certainly required to confirm our preliminary results which suggest that lead previously stored in the organism can be mobilized during pregnancy and hence transferred to the foetus. Experiments involving more animals are under way but we thought useful to already report the preliminary results to stimulate further investigation on the assessment of the potential risk of lead for the foetus. References Buchet J P , Roels H , Hubermont G , Lauwerys R : Effect of lead on some parameters of the heme biosynthetic pathway in rat tissues in vivo Toxicology 6, 21-34 (1976) Eisenstein R , Kawanoue S : The lead line in bone A lesion apparently due to chondroclastic indigestion Amer J Pathol 80, 309-314 (1975) Hsu F S , Krook L , Shively J N , Duncan J R , Pond W G : Lead inclusion bodies in osteoclasts. Science 181, 447-448 (1973) Hubermont G , Buchet J P , Roels H , Lauwerys R : Effect of short-term administration of lead to pregnant rats Toxicology 5, 379-384 (1976) Van Mullem P H , Stadhouders A M : Bone marking and lead intoxication Early pathological changes in osteoclasts Virchows Arch 15, 345-350 (1974)
Received March 28, 1977 / Accepted June 6, 1977
Int Arch Occup Environ Hlth 40, 33-36 (1977)
Culutlional aNul Eriumnental (k
Health
©)Springer-Verlag
1977
Mobilization of Lead During Pregnancy in Rats* J.P Buchet, R Lauwerys, H Roels and G Hubermont Unite de Toxicologie Industrielle et Mdicale, University of Louvain, 30 Clos Chapelle-aux-Champs, B-1200 Brussels, Belgium
Summary Lead was administered to three groups of 4 weeks-old female rats at the dose of 1 ppm in drinking water The first group received lead continuously for 150 days before mating (with non-lead-treated male rats), during pregnancy and during 3 weeks after delivery In the second group lead was also administered for 150 days before mating but was withdrawn during pregnancy and during the post natal period In the third group lead was administered for 150 days; the animals were then kept without lead exposure during the 50 days before making them pregnant, during pregnancy and during the post natal period A control group received only demineralized water. On day 21 after delivery the mothers and their newborns were sacrificed and various parameters of blood-lead concentration (Pb-B), hematocrit (Htc), hemoglobin (Hb), free erythrocyte porphyrins concentration (FEP), 6-aminolevulinate dehydratase activity (ALAD) and tissues ALAD, free tissue porphyrins (FTP), lead concentration (Pb-T) were determined. In mothers and in newborns lead concentration in blood and in soft tissues were increased over control values only in the group in which lead had been stopped for 50 days before pregnancy This lead accumulation in soft tissues was associated with an increased FTP in liver, heart and brain of the mothers and in liver of the newborns This observation suggests that lead stored in the organism can be mobilized during pregnancy. Key words: Lead in blood Lead in tissues ALAD in blood ALAD in tissues Porphyrins in blood Porphyrins in tissues Transplacental transfer We have previously studied the effect of short-term oral administration of lead on various parameters of the heme biosynthetic pathway in pregnant rats and their offspring (Hubermont et al 1976) Three doses of lead ( 10, 1 and 0 1 ppm in drinking water) were administered for 3 weeks before mating, during pregnancy and during * Supportedbyagrantfrom the Commission of the European Communities (Project no 035-74-1Env B) and the Fonds de la Recherche Scientifique M 6dicale (Belgium)
34
J P Buchet et al.
three weeks after delivery No significant biological changes (except increased lead mothers concentration in blood and in kidney in the 10 ppm group) were found in the (ALAD) dehydratase -aminolevulinate 6 group ppm 10 but in the newborns of the porphyrins activity in blood and in kidney was significantly decreased and free tissue values. control over increased significantly was kidney (FTP) concentration of We thus concluded that the developing organism is more susceptible to the biothe logical action of lead than is the organism of adult animals and on the basis of preduring administered lead of level no-effect the biochemical parameters studied whether tested now have We ppm 1 around is gnancy and in the neonatal period 1 ( ppm) to female rats before longterm (150 and 200 days) administration of lead level found during the short-term ,no-effect" the making them pregnant will modify of the animal could be mobilized life early during study and whether lead administered pregnancy. subsequent a and transferred to the foetus during Four weeks-old female Sprague-Dawley rats were used Lead was administered as Pb(NO 3 ) 2 at the dose of 1 ppm in demineralized drinking water There were 4 groups of 8 female rats: the control group (a) received only demineralized water, 4 animals during 150 days and 4 during 200 days; since no significant difference in the biological measurements was found between the two subgroups, the results were pooled for purpose of comparison with groups (b), (c) and (d); group (b) received lead continuously for 150 days before mating (with non-lead-treated male rats), during for pregnancy and during 3 weeks after delivery; in group (c) lead was administered post the during and pregnancy during withdrawn 150 days before mating and was then natal period; in group (d) lead was administered for 150 days; the animals were during pregnant, them making before days 50 the kept without lead exposure during pregnancy and during the post natal period At time of sacrifice the animals in this group were thus 50 days older than those of groups (b) and (c). Before mating the Pb-B levels of the animals in group (d) were determined. Not all the animals became pregnant and in order to have a similar number of animals in each group only 4 pregnant animals were examined in each group At delivery, only 8 pups per mother were kept alive On day 21 after delivery the animals were homosacrificed Blood collections, whole body perfusion and preparation of tissue tissues and blood The 1976) al et (Buchet previously genates have been described of 3 pups were pooled to obtain sufficient sample volume for all the determinations. beFor each mother two ,,newborn" values were thus obtained which were averaged whole on performed were analyses biochemical following The fore statistical analysis blood: ALAD activity (in nmoles -ALA transformed/min x ml erythrocytes), hematoml crit (expressed in %), free erythrocyte porphyrin (FEP) concentration (in g/ 100 conand erythrocytes), hemoglobin (Hb) concentration (in g/100 ml whole blood) On the centration of lead in blood (Pb-B) (expressed in gg lead/100 ml whole blood) measurements whole homogenate of liver, kidney, heart and brain, the following mg protein), were performed: ALAD activity (in nmoles 6-ALA transformed/min x (expressed (Pb-T) tissue FTP concentration (in ng/mg protein), concentration of lead in methods analytical in ng lead/mg protein) and concentration of tissue protein The used have been described previously (Buchet et al , 1976). older The treatments and the slight age difference between the groups (group d mothers the of weight average than groups b and c) had no significant effect on the
35
Mobilization of Lead During Pregnancy in Rats
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at sacrifice (individual values ranging from 290 to 320 g) and of their pups (individual values ranging from 38 to 48 g). In mothers lead concentration in blood and in tissues was lower in the groups receiving lead continuously (b) or up to the beginning of pregnancy (c) than in the group (d) in which lead had been stopped for 50 days before pregnancy (Table 1).
36
J P Buchet et al.
In this group lead concentration in blood and kidney was significantly increased over control value This observation suggests that lead stored in the organism can be mobilized during pregnancy. This hypothesis is reinforced by the fact that just before mating the Pb-B of the female rats of group (d) were in the normal range (< 3 g/ 100 ml whole blood). In the same maternal group (d) FTP in liver, heart and brain were significantly increased (Table 1) but ALAD was not inhibited in blood or in tissue There was no effect of lead treatment on Htc, Hb and FEP level. In newborns, the change in lead concentration of blood and tissues paralleled that found in mothers (Tab 1) A significant increase was found in blood, liver, kidney, heart and brain of newborns of group (d), i e , that in which Pb-administration to the mothers was stopped 50 days before pregnancy (One way analysis of variance p < 0 05). The increased lead levels found in group (d) were not associated with a reduction of ALAD in blood or tissues but FTP of liver were significantly increased (p < 0 05, Table 1) Like in mother, no change in Htc, Hb and FEP level were found. In summary, the most significant observation made during this preliminary investigation is the increased accumulation of lead in newborns of mothers which had received 1 ppm Pb for 150 days and had been on a lead free regime for 50 days before pregnancy (group d) This lead accumulation was associated with an increased FTP concentration in liver of newborns It is surprising that this phenomenon was not observed when 1 ppm lead was administered continuously up to the beginning of gestation (group b) or throughout pregnancy (group c) since the lead body burden of these animals was certainly higher than that of group (d) This could be due to a reduction of the osteocytic or osteoclastic resorption of bone in both groups (b) and (c) (Eisenstein and Kawanoue, 1975 ; Hsu et al 1973) Lead inclusions have been described in osteoclast (Hsu et al , 1973 ; Von Mullem and Stadhouders , 1974) This could be accompanied by a reduction of their metabolic activity In group (d), the reduction of lead concentration in bone during the 50 day-period without lead administration before mating might have been sufficient to restore the osteoclastic activity. Further informations are certainly required to confirm our preliminary results which suggest that lead previously stored in the organism can be mobilized during pregnancy and hence transferred to the foetus. Experiments involving more animals are under way but we thought useful to already report the preliminary results to stimulate further investigation on the assessment of the potential risk of lead for the foetus. References Buchet J P , Roels H , Hubermont G , Lauwerys R : Effect of lead on some parameters of the heme biosynthetic pathway in rat tissues in vivo Toxicology 6, 21-34 (1976) Eisenstein R , Kawanoue S : The lead line in bone A lesion apparently due to chondroclastic indigestion Amer J Pathol 80, 309-314 (1975) Hsu F S , Krook L , Shively J N , Duncan J R , Pond W G : Lead inclusion bodies in osteoclasts. Science 181, 447-448 (1973) Hubermont G , Buchet J P , Roels H , Lauwerys R : Effect of short-term administration of lead to pregnant rats Toxicology 5, 379-384 (1976) Van Mullem P H , Stadhouders A M : Bone marking and lead intoxication Early pathological changes in osteoclasts Virchows Arch 15, 345-350 (1974)
Received March 28, 1977 / Accepted June 6, 1977
