Zinc supplementation of pregnant rats with adequate zinc nutriture suppresses immune functions in their offspring.
ABSTRACT The knowledge about consequences of marginal zinc (Zn) deficiency and Zn supplementation during pregnancy on immune function in the offspring is limited. The aim of this study was to examine whether effects of mild Zn deficiency and subsequent Zn supplementation during pregnancy persist after weaning and affect immune function of the offspring. Adult female rats were fed a Zn-adequate diet (ZC, n = 8) or a Zn-deficient diet (ZD, n = 8) from preconception through lactation. Pregnant rats were supplemented with either Zn (1.5 mg Zn in water) or placebo (water) 3 times/wk throughout pregnancy. Pups were orally immunized with cholera toxin and bovine serum albumin-dinitrophenol (DNP) 3 times at weekly intervals and killed 1 wk after the last dose. Proliferation and cytokine responses in lymphocytes from Payer's patches and spleen, and antigen specific antibodies in serum were studied. Zn supplementation of ZD dams led to enhanced lymphocyte proliferation and IFN-gamma responses in pups ZDZ+. In contrast, Zn supplementation of ZC dams suppressed these responses in pups ZCZ+. Total and DNP-specific IgA responses were lower in pups of the Zn-deficient group compared with the Zn-adequate group. Relative thymus weight was greater in the pups (ZDZ-) of ZD placebo-supplemented dams compared with the other groups at 31 d of age. Prepregnancy and early in utero Zn deficiency affected IgA responses in pups that could not be restored with Zn supplementation during pregnancy. Zn supplementation of ZC dams induced immunosuppressive effects in utero that may also be mediated through milk and persist in the offspring after weaning.
- SourceAvailable from: Shannon Kelleher[Show abstract] [Hide abstract]
ABSTRACT: Dietary analysis predicts that marginal Zn deficiency is common in women of reproductive age. The lack of reliable biomarkers limits the capacity to assess Zn status and consequently understand effects of maternal Zn deficiency. We determined effects of marginal maternal Zn deficiency on mammary gland function, milk secretion, and milk composition in mice. Mice (n = 12/diet) were fed marginal (ZD; 15 mg Zn/kg diet) or adequate (ZA; 30 mg Zn/kg diet) Zn diets for 30 d prior to conception through mid-lactation. Mice fed the ZD had a higher plasma Zn concentration (~20%; P < 0.05) but lower milk Zn concentration (~15%; P < 0.05) compared with mice fed the ZA. ZnT2 abundance was higher (P < 0.05) in mice fed the ZD compared with mice fed the ZA; no effect on ZnT4 abundance was detected. The Zn concentration of mammary gland mitochondria tended to be ~40% greater in mice fed ZD (P = 0.07); this was associated with apoptosis and lower milk secretion (~80%; P < 0.01). Total milk protein was ~25% higher (P < 0.05), although the abundance of the major milk proteins (caseins and whey acidic protein) was lower (P < 0.05) in mice fed the ZD. Proteomic analysis of milk proteins revealed an increase (P < 0.05) in four proteins in mice fed the ZD. These findings illustrate that marginal maternal Zn deficiency compromises mammary gland function and milk secretion and alters milk composition. This suggests that lactating women who consume inadequate Zn may not produce and/or secrete an adequate amount of high quality milk to provide optimal nutrition to their developing infant.Journal of Nutrition 02/2012; 142(4):655-60. · 4.20 Impact Factor
- 02/2012; , ISBN: 978-953-51-0138-3
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ABSTRACT: Anemia and iron deficiency affect billions of people worldwide, especially women of reproductive age, pregnant women, and young children. Many countries have iron and folic acid supplementation programs for pregnant women. However, the impact of these programs is uncertain. Multiple-micronutrient supplementation during pregnancy has been advocated; however, it is unclear whether this has additional advantages. Overall, programs have shown only modest impact on increasing birth weight. This review discusses the present state of knowledge on interventions to improve iron status during pregnancy and reproductive health, and investigates other possibilities such as supplementation prior to conception to improve maternal and child health.Nutrition Reviews 11/2011; 69 Suppl 1:S78-86. · 4.60 Impact Factor
The Journal of Nutrition
Zinc Supplementation of Pregnant Rats with
Adequate Zinc Nutriture Suppresses Immune
Functions in Their Offspring1
Rubhana Raqib,2* Mohammad Bakhtiar Hossain,2,3Shannon L. Kelleher,3Charles B. Stephensen,3,4
and Bo Lo ¨nnerdal3
International Centre for Diarrhoeal Diseases Research, Bangladesh (ICDDR,B), Dhaka-1212, Bangladesh;3Department of Nutrition,
University of California, and4USDA Western Human Nutrition Research Center, Davis, CA 95616
The knowledge about consequences of marginal zinc (Zn) deficiency and Zn supplementation during pregnancy on
immunefunctionintheoffspringislimited.The aimofthis studywastoexaminewhethereffectsofmildZndeficiencyand
subsequent Zn supplementation during pregnancy persist after weaning and affect immune function of the offspring.
Adult female rats were fed a Zn-adequate diet (ZC, n ¼ 8) or a Zn-deficient diet (ZD, n ¼ 8) from preconception through
lactation. Pregnant rats were supplemented with either Zn (1.5 mg Zn in water) or placebo (water) 3 times/wk throughout
pregnancy. Pups were orally immunized with cholera toxin and bovine serum albumin-dinitrophenol (DNP) 3 times at
weekly intervals and killed 1 wk after the last dose. Proliferation and cytokine responses in lymphocytes from Payer’s
patches and spleen, and antigen specific antibodies in serum were studied. Zn supplementation of ZD dams led to
enhanced lymphocyte proliferation and IFN-g responses in pups ZDZ1. In contrast, Zn supplementation of ZC dams
suppressed these responses in pups ZCZ1. Total and DNP-specific IgA responses were lower in pups of the Zn-deficient
group compared with the Zn-adequate group. Relative thymus weight was greater in the pups (ZDZ2) of ZD placebo-
supplemented dams compared with the other groups at 31 d of age. Prepregnancy and early in utero Zn deficiency
affected IgA responses in pups that could not be restored with Zn supplementation during pregnancy. Zn supplementation
of ZC dams induced immunosuppressive effects in utero that may also be mediated through milk and persist in the
offspring after weaning.J. Nutr. 137: 1037–1042, 2007.
Observational and experimental studies have highlighted the
importance of zinc (Zn)5status in immune function as illustrated
Severe Zn deficiency has been associated with thymic and spleen
atrophy, impaired T cell-mediated responses, and increased
host to replenish peripheral lymphocytes, causing increased
susceptibility to infectious diseases (2). cDNA array analysis of
thymus from moderately Zn-deficient mice demonstrated that
aberrations in specific thymic mRNA take place long before T
lymphocyte changes are detectable by Florescent Associated Cell
Sorter analysis (3). Impaired immune response to vaccination in
experimental animals and in elderly patients with Zn deficiency
has been reported (4–7), although the findings are not always
Moderate to severe Zn deficiency during pregnancy in
experimental animals has been related to adverse effects on
offspring, including high rates of fetal resorption, reduced litter
size, congenital malformations, reduced splenocyte responsive-
ness to mitogen (11–13), and reduced serum levels of IgG2a and
marginallyreduce the immunodeficiency, but the defect persisted
to a milder extent in the F2 and F3 progenies (14,15). Observa-
on adverse fetal outcome relating to Zn deficiency, most likely
of the effect of maternal Zn deficiency on the infant’s immune
status are scarce. One study of a 6-mo follow-up of infants in
Bangladesh demonstrated that only low birth weight (but not
normal birth weight) infants of mothers who received Zn
supplementation during pregnancy had reduced risk of diarrhea,
dysentery, and impetigo compared with the placebo group (16).
Another study by the same group documented the benefit to
1Supported by the Ellison Medical Foundation at the Department of Nutrition,
University of California Davis.
5Abbreviations used: AAS, atomic absorption spectrophotometry; BSA, bovine
serum albumin; Con A, Concanavalin A; CT, cholera toxin; DNP, dinitrophenol;
PP, Peyer’s patch; ZC, control dams that received adequate zinc in the diet;
ZCZ1, pups of control dams that received zinc supplementation; ZCZ2, pups of
control dams that received placebo; ZD, zinc-deficient dams that received diet
marginally deficient in zinc; ZDZ1, pups of zinc-deficient dams that received zinc
supplementation; ZDZ2, pups of zinc-deficient dams that received placebo; Zn,
* To whom correspondence should be addressed. E-mail: email@example.com.
0022-3166/07 $8.00 ª 2007 American Society for Nutrition.
Manuscript received 25 October 2006. Initial review completed 30 November 2006. Revision accepted 23 January 2007.
by guest on May 30, 2013
infant immune status by showing reduction in infectious and
diarrheal disease morbidity incidences throughout the 1st period
of life (17). However, direct investigations of the effects of Zn
deficiency and supplementation during pregnancy on immune
function in offspring have not been conducted.
weaning and can be corrected by Zn supplementation during
pregnancy. Mild Zn deficiency is more common in human
populations. Therefore, we investigated effects of mild Zn
deficiency during pregnancy on the development of the immune
system in offspring using a rat model. Public health programs of
prenatal Zn supplementation do not take into account maternal
Zn status, which is an unexplored area. Thus, we further exam-
on the ability to boost immune response in the offspring after
Materials and Methods
Rats. The study complied with the Guide for the Use and Care of
Laboratory Rats and was conducted under the auspices of Animal
Resource Services of the University of California, Davis, which is
accredited by the American Association for the Accreditation of Labo-
ratory Animal Care. Virgin female Sprague-Dawley rats (n ¼ 16; 7–8 wk
in stainless steel hanging cages under constant conditions (22?C, 65%
humidity) with a 12-h-dark:light cycle and consumed food ad libitum.
After consumption of standard nonpurified rat diet (LabDiet) for a 3-d
acclimatization period, rats were randomly assigned to 1 of 2 experi-
mental diets and allowed to consume the food and deionized water ad
libitum. We developed the Zn-deficient pregnant rat model, as described
previously (18). The Zn-adequate control group (ZC; n ¼ 8) was given a
diet containing 25 mg Zn/g diet and the other group of rats (ZD; n ¼ 8)
received a diet marginally deficient in Zn (7 mg Zn/g diet). Rats received
postparturition. Following pregnancy confirmation, these rats were
furtherdividedinto2 subgroupstoreceive either Zn(1.5 mgZn inwater,
status in pregnant dams was determined by measuring Zn in serum
collected from tail blood (18).
After birth, different groups of pups were designated as follows:
ZCZ1, pups of control dams that received Zn supplementation; ZCZ2,
pups of control dams that received placebo; ZDZ1, pups of Zn-deficient
dams that received Zn supplementation; ZDZ2, pups of Zn-deficient
dams that received placebo. At birth, litter size was recorded, but body
weight was not measured. Pups of ZD dams were much smaller than
those of the ZC group. Postnatal survival was less in the ZD group. Pups
were culled to 8 pups/dam at d 3. After weaning, pups received the same
diet as their mothers. Pups were killed at d 31 to determine the long-term
effects of maternal Zn intake during pregnancy on cellular and mucosal
immune function in the offspring. Body weight was first measured at d
10 and thereafter every week. Rats were killed by asphyxiation with
CO2, body weight was measured, and blood was collected immediately.
Immunization. We immunized pups from 4 treatment groups (n ¼ 12 3
4) via oral gavage at d 10, 17, and 24 with cholera toxin (CT; 10 mg/
immunization) and T lymphocyte-dependent antigen, dinitrophenol-
bovine serum albumin (DNP-BSA; 50 mg/immunization) in a total
volume of 0.1 mL 3% sodium bicarbonate buffer to neutralize gastric
acid (Fig. 1). Bicarbonate only wasgiven to pups (n ¼ 4 3 4) as a control.
Tissue and blood collection. We recorded the weight of thymus. Cells
were harvested from spleen and single cell suspensions were made by
were then used for cell culture and the proliferation assay. Peyer’s patch
(PP) lymphocytes were tweezed out from the small intestine, applied to a
grinder, and passed through nylon mesh to obtain a single cell suspension
(19). Samples of intestine and liver were dissected and immediately snap-
frozen in liquid nitrogen and frozen at 280?C for Zn analysis. Blood
obtained by cardiac puncture was collected into trace element-free vials
Mineral analysis. Tissue minerals were measured by atomic absorption
spectrophotometry, as described previously (20). Briefly, ;0.2 g of
tissues was digested in acid-washed vials with 3.0 mL of 16 mol/LHNO3
for 48 h. Samples were boiled to a volume of ;1.0 mL and ultra-pure
water added to a volume of 5.0 mL. Serum (120 mL) was digested in 1.0
mL of 1 mol/L HNO3for 48 h prior to absorption spectrophotometry.
Lymphocyte proliferation response. Mononuclear cells were isolated
from spleen and PP in the small intestine and single-cell suspensions were
cultured, as previously described (21), using 3 doses of CTand 3 doses of
2.0 mg/L) of the mitogen concanavalin A (Con A) was used as control
stored at 270?C until used. Proliferation was assessed by bromodeoxy-
uridine incorporation using standard methods and commercial reagents
(US Biochemicals). Data were expressed as absorbance at 450 nm
(reference wavelength, 690 nm).
cell type 2, IL-4 and IL-10, were measured in the culture supernatant of
measured cytokines using commercial enzyme immunoassays (R&D
Systems). The lowest limit of detection was 10 ng/L for IFN-g and 1 ng/L
for IL-4 and IL-10.
Antigen-specific antibody responses in serum. DNPandCT-specific
IgG and IgA, as well as total IgA antibodies, were measured in serum
according to the method described previously (22). In brief, polystyrene
microtiter plates (Nunc-Maxisorp) were coated with DNP-ovalbumin in
carbonate buffer (0.1 mol/L sodium bicarbonate and 5 mmol/L magne-
sium chloride, pH 9.8) and incubated overnight at 4?C and the standard
fed Zn-restricted (7 mg Zn/g diet) or Zn-adequate diet (25 mg Zn/g diet) and were
supplemented with either Zn (1.5 mg Zn in water, 3 times/wk) or placebo
(water). Pups were immunized via oral gavage at d 10, 17, and 24 with CT (10
mg/immunization) and DNP-BSA (50 mg/immunization) in 0.1 mL of 3% sodium
bicarbonate buffer. Only bicarbonate was given to pups as a control.
Trial profile demonstrating number of dams in each group that were
1038 Raqib et al.
by guest on May 30, 2013
procedure was followed. Data were expressed as mg/L of antibodies.CT-
specific responses were measured, as previously described (21).
Statistical analyses. Statistical analyses were conducted using the
statisticalsoftwarepackagesSIGMASTAT (version3.1; JandelScientific)
and SPSS for WINDOWS (release 10; SPSS Institute). When a variable
was not normally distributed, an appropriate transformation (e.g. log or
square root) was used to better achieve approximate normality. Analyses
were performed on the transformed variables to meet the underlying
assumptions of the statistical tests used. When the data could not be
normalized, nonparametric analysis (rank-sum test) was performed.
Differences were significant at P , 0.05. Results were expressed as
means 6 SE. Data were analyzed using 2-way ANOVA to test for main
effects of dietary Zn content and Zn supplementation and their inter-
action during pregnancy. When the 2-way interaction was significant
(P , 0.05), 1-way ANOVA was conducted, followed by Tukey’s test.
Repeated measures ANOVA was used to test body weight data. Body
weights on d 10, 24, and 31 were used as within-subjectvariable with Zn
supplementation as the between-subject factor.
Body and thymus weights. Body weight in the ZCZ1 group
on d 10 was higher (P ¼ 0.001) than in the ZCZ group, but body
weights on d 24 and d 31 did not differ (data not shown). Body
weight in the ZDZ1 group was higher (P ¼ 0.001) on d 10 than
in the ZDZ2 group. Gain in body weight over the study period
was higher in the pups of the ZD groups than in the ZC groups
(P ¼ 0.03) (data not shown).
Thymus weight was significantly higher in the ZDZ2 group
than in the ZDZ1 group, but it did not differ between the 2 ZC
groups (data not shown). The weight of the thymus in the
ZDZ2 group wasmarkedly higher than those in the ZCZ2(P ¼
0.001) and ZCZ1 (P ¼ 0.002) groups. When corrected for body
weight, the thymus weight in the ZDZ2 group was higher than
in the ZDZ1 and the ZCZ2 groups and tended to be higher
than the ZCZ1 group (P ¼ 0.053) (Fig. 2).
Zn concentrations. At the preconception stage, serum Zn
Zn in serum, liver, and intestinal tissue compared with the ZD
groups. However, within the ZC and ZD groups, Zn supplemen-
tation did not affect serum or tissue Zn concentrations (Table 1).
Lymphocyte proliferation response. The optimum dose for
stimulation with Con A was 1.0 mg/L, although significant
differences between the groups were obtained at 0.5 mg/L.
Spleen lymphocytes (splenocytes) in the ZCZ1 group had a
lower response compared with the ZCZ2 group (P ¼ 0.005) at
0.5 mg/L Con A and tended to be lower at 2.0 mg/L (P ¼ 0.06)
(Table 2). However, the ZDZ2 and the ZDZ1 groups did not
differ at any dose of the mitogen.
PP lymphocytes of the ZDZ1 group tended to have a greater
shown) (P ¼ 0.053). However, the proliferation response of PP
Proliferation responses of splenocytes or PP lymphocytes to
the immunogens CT or BSA were below the detection limit in
most rats (data not shown).
Cytokines in cultured lymphocyte supernatant. Cytokines
IL-4 and IL-10 were detectable in only a few rats (data not
shown); however, high levels of IFN-g were obtained in the
supernatant. Within the ZD and ZC groups, the levels of IFN-g
in splenocyte cultures were not affected by Zn supplementation.
However, IFN-g levels were higher in the ZDZ1 group than in
the ZCZ1 group (P ¼ 0.01) (Fig. 3). This may reflect a
suppressive effect of additional Zn in the ZCZ1 group in the
release of IFN-g in the culture supernatant. The IFN-g levels did
not differ between the ZDZ2 and ZCZ2 groups.
In the cultures of PP lymphocytes, low levels of IFN-g were
detected in the supernatant. The IFN-g levels did not differ
between the ZD and ZC groups (data not shown).
Antigen-specific responses. Levels of antibody against CTor
BSA were very low and in most cases were below the detection
limit (data not shown). However, DNP-specific IgA and IgG1
were measurable in serum.
Total IgA concentrations were higher in pups in the ZCZ2
group compared with the ZCZ1 group (Table 3). However, the
deficient dams supplemented with Zn or placebo. Values represent means 6
SEM, n ¼ 12. Means without a common letter differ, P , 0.05.
Relative thymus weights of 31-d-old pups of Zn-adequate and Zn-
Concentrations of Zn in serum of Zn-adequate and Zn-deficient dams at preconception and in serum and tissues of pups
of Zn-adequate and Zn-deficient dams supplemented with Zn or placebo1
Serum, mmol/L1.66 6 0.03 1.69 6 0.06 2-way ANOVA (P-values)
Dietary ZnZn supplementInteraction
Pups, 31 d old
1.65 6 0.06
24.5 6 0.64
26.3 6 0.63
1.53 6 0.09
23.6 6 0.68
24.9 6 0.97
0.94 6 0.06
20.4 6 0.57
23.9 6 0.64
0.79 6 0.05
18.7 6 0.59
22.6 6 1.24
1Data are means 6 SE; n ¼ 12 (pups) or 8 (dams).
Excess zinc suppresses immunity in offspring 1039
by guest on May 30, 2013
IgA (P ¼ 0.01) as well as DNP-specific IgA (P ¼ 0.04) compared
with those in the ZD groups. Total IgA levels in the ZCZ2 group
weresignificantlyhigherthan theothergroups(Table3).Levels of
DNP-specific IgG1 did not differ among the groups.
Direct benefits in infant immune status as a result of maternal Zn
supplementation have shown reductions in infectious and
diarrheal disease morbidity incidences throughout the 1st period
of life (17). However, direct investigations of the effects of Zn
deficiency and supplementation during pregnancy on immune
function in offspring have not been evaluated. Here, we report
that Zn deficiency during pregnancy in rats was reflected by
changes in immune function of the offspring such that Zn
supplementation to Zn-deficient dams restored cellular immune
deficits in the offspring but could not restore humoral immunity.
Moreover, Zn supplementation to dams with adequate Zn
nutriture lead to suppressed cellular immunity in their pups,
suggesting that Zn ‘‘over-nutrition’’ may be as important as Zn
‘‘under-nutrition’’ in vulnerable populations.
The immunological consequences of severe Zn deficiency
with reductions in size of spleen and thymus, leading to cellular
immune defects (14,23,24) and lower antigen-specific, antibody-
mediated responses in offspring (25).Zn supplementation of Zn-
deficient animals and humans restores immune deficiency,
including lymphocyte function, delayed hypersensitivity re-
sponse, and cell-mediated immune responses (26,27). However,
studies directly investigating effects of Zn supplementation
during pregnancy on immune function in their infants or older
deficiency, observational studies of Zn supplementation of
pregnant women have generated conflicting results. One study
showed high IgG levels in cord blood of mothers supplemented
to Hib-conjugate vaccine in infants of mothers supplemented
with Zn; however, fewer anergic responses to delayed hypersen-
infants after maternal Zn supplementation (17,28), suggesting a
positive effect of Zn supplementation only in potentially Zn-
deficient infants. Inthis study,we produced mildZn deficiency in
pregnant dams. This condition is relatively common in human
supplementation of Zn-deficient dams led to increased prolifer-
withtheplacebogroup.SecretionofIFN-g inresponsetoaT cell
mitogen, con A, was higher in the young pups from Zn-
supplemented, Zn-deficient dams compared with the offspring
of Zn-supplemented, Zn-adequate control dams. These findings
suggest that repletion of Zn status during pregnancy restores the
mucosal cellular immune function of offspring and thus has a
beneficial effect on the immune system of the progeny.
Zn supplementation has been shown to reverse immune
have been documented to impair T cell function (26,29) and
block IFN-a production (30). Only 1 study in elderly humans
reported depressed immune function after oral intake of high Zn
(31). Reports on the effect of surplus Zn during Zn-adequate
pregnancies on human infants are absent and limited in exper-
imental animals. One study in adult female mink reported that
alopecia, lymphopenia, suppressed lymphocyte proliferation
response, and reduced growth rate in their offspring (32). The
IFN-g production in the offspring, suggesting inhibitory or
suppressive effects of excess Zn on T lymphocyte functions. The
level of Zn supplementation used in our study was very modest
and similar on a body weight basis to amounts that pregnant
trations and are more susceptible to increased Zn levels than
other cells (26). High Zn concentration suppressed alloreactivity
in mixed lymphocyte culture (33), decreased lymphocyte reac-
tivity to mitogens (34,35), and blocked IFN-a production (30).
Lymphocyte proliferation response (bromodeoxyuridine) in splenocytes from pups of
Zn-adequate and Zn-deficient dams supplemented with Zn or placebo1
2-way ANOVA (P-values)
Con A ZCZ2
Dietary Zn Zn supplementInteraction
0.42 6 0.03a
0.82 6 0.09
0.93 6 0.10
0.30 6 0.05b
0.70 6 0.09
0.69 6 0.07
0.32 6 0.04b
0.63 6 0.09
0.70 6 0.08
0.27 6 0.02b
0.78 6 0.09
0.90 6 0.11
1Data are mean absorbances 6 SE, n ¼ 12. Means in a row with superscripts without a common letter differ, P , 0.05.
splenocytes from pups of Zn-adequate and Zn-deficient dams supplemented
with Zn or placebo. Values represent means 6 SEM; n ¼ 12. Means without a
common letter differ, P , 0.05.
Concentration of IFN-g in culture supernatants of Con A-stimulated
1040 Raqib et al.
by guest on May 30, 2013
The inhibitory effects of excess Zn may also result from Zn-
homeostasis (38), because both copper and vitamin A are im-
portant for T cell function. The findings indicate that excessive
Zn supplementation during pregnancy had an adverse carryover
effect on the offspring.
Zn deficiency leads to decreased humoral response (1,27) and
Zn supplementation restores antibody response. In a study of
postpartum marginal Zn deprivation in mouse dams, the ability
to mount antibody-mediated responses in suckling pups was
reduced (25). Effects of Zn deficiency in dams on the immune
responses of pups mediated via milk were reversed by Zn
supplementation of the pups. However, in our study, prepreg-
nancy Zn deficiency in rat dams caused a permanent defect
imprinted in utero in the humoral arm of immunity that could
not be restored even after Zn supplementation to the pregnant
dams. Indeed, lowered response to B cell mitogen was reported
in the offspring of marginally Zn-deficient monkey dams (39).
One drawback of this study was that oral immunization of
suckling rats started at the age of 10 d and given at weekly
intervals when the immune system of these pups was still
immature (21). The lack of antibody response or lymphocyte
proliferation response to specific antigens, CTor BSA, might be
due to the immaturity of the mucosal immune system or a result
of induced tolerance due to short intervals of immunization.
An interesting observation in this study was the increased size
of the thymus in the 31-d-old offspring of ZD dams receiving
placebo. Studies in experimental Zn-deficient models have
shown that severe Zn deficiency causes thymic atrophy and
decreases thymic size (2,27). However, to our knowledge, this is
the 1st report to show that mild Zn deficiency during fetal
development and continued suboptimal Zn nutriture during
infancy resulted in enlarged thymic size in the offspring. Despite
having smaller size at birth, a common feature during maternal
Zn deficiency in animals (14,24), these young Zn-deficient pups
also had higher weight gain by adolescence. The difference in
thymus weights remained significant after being corrected for
body weight. In humans, thymus size is largest at puberty and is
reduced at adulthood to the size at birth. It is possible that the
ZDZ2 group did not attain the natural size of thymus by
adolescence due to delayed maturity. It is also possible that
marginally Zn-deficient pups being kept in a relatively sterile
environment and nutritional affluence might have had a rapid
catch-up growth causing obesity in adult life (40,41).
In conclusion, we foundthateffects of maternal Zndeficiency
and humoral immune functions of young pups. In addition, Zn
supplementation to Zn-deficient dams during pregnancy cor-
rected cellular immune function, whereas that to Zn-adequate
dams was immunosuppressive and persisted in the offspring in
later life. Humoral immunodeficiency in pups could not be
restored with maternal Zn supplementation. Thus, in the public
health context, further research assessing optimum dosage of
Zn supplementation during pregnancy, especially linking it to
immunological outcome in infants and children, is crucial.
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dams supplemented with Zn or placebo1,2
Two-way ANOVA (P-values)
Dietary ZnZn supplement Interaction
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23.3 6 8
45 6 10
463 6 220b
16.6 6 13
166 6 70
27.3 6 12b
6 6 2.4
29 6 4
47.1 6 13b
4 6 1
36 6 12
1Data are means 6 SE, n ¼ 12. Means in a row with superscripts without a common letter differ, P , 0.05.
2Pups were orally immunized with DNP-BSA and CT.
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1042Raqib et al.
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