Gestational exposure to mercury leads to persistent changes in T-cell phenotype and function in adult DBF₁ mice

Figures & data

Table 1.  Litter statistics from 3 independent timed-breed experiments to evaluate the persistent immune effects of gestational exposure to 10 ppm HgCl₂.

	Experiment 1	Experiment 2	Experiment 3
Control
Number of Litters	2	2	2
Total pups	6	8	12
Fetal deaths	0	0	0
Number of males	3	2	6
Average weight of males (g ± SD)	23.8 ± 0.80	22.81 ± 3.42	26.19 ± 5.11
Number of females	3	6	6
Average weight of females (g ± SD)	22.57 ± 0.47	20.18 ± 1.97	25.10 ± 2.30
Mercury-Exposed
Number of Litters	3	2	2
Total pups	12	7	4
Fetal deaths	0	0	0
Number of males	6	4	1
Average weight of males (g ± SD)	24.07 ± 1.38	23.91 ± 2.70	22.10 ± 0.54
Number of females	6	3	3
Average weight of females (g ± SD)	22.80 ± 0.50	21.04 ± 1.12	20.60 ± 0.18

Figure 1.  The effects of prenatal HgCl₂ exposure on thymic and splenic cellularity. Cell numbers in the (A) thymus and (B) spleen of 10-wk-old DBF₁ mice exposed to plain water (control) or 10 ppm mercuric chloride during gestation were enumerated. Data were collected from six control mice (3 female and 3 male DBF₁ mice) and 12 mercury-exposed mice (6 female and 6 male DBF₁ mice). The differences between mercury-exposed mice and sex-matched control mice were considered significant when p < 0.05.

Table 2.  The effect of prenatal mercuric chloride exposure on CD4/CD8-expressing cells in the thymus.

	CD4^+CD8^−	DP	DN	CD8^+CD4^−	CD4^+/CD8^+
Control male	9.29 ± 1.05	77.94 ± 1.86	6.04 ± 1.32	6.83 ± 0.64	1.42 ± 0.22
Mercury male	8.37 ± 0.41	80.59 ± 1.26	4.50 ± 0.50	6.54 ± 1.00	1.90 ± 0.23
Control female	8.43 ± 0.66	77.58 ± 1.75	6.44 ± 1.44	7.55 ± 1.12	1.62 ± 0.70
Mercury female	6.92 ± 0.71	80.61 ± 1.13	6.43 ± 1.37	6.05 ± 0.84	1.26 ± 0.24

The percentages in each phenotypic compartment were determined by flow cytometry and expressed as mean ± SEM. The data presented here from 22 control mice (14 female and 8 male DBF₁ mice) and 21 mercury-exposed mice (12 female and 9 male DBF₁ mice) from three independent experiments.

Table 3.  The effect of prenatal mercuric chloride exposure on CD4/25-expressing cells in the thymus.

	CD4^+CD25^+
	Percentage of cells	Number of cells (×10^5)
Control Male	1.30 ± 0.12	3.30 ± 0.40
Mercury Male	1.36 ± 0.12	3.66 ± 0.39
Control Female	1.26 ± 0.12	3.41 ± 0.31
Mercury Female	1.17 ± 0.13	2.91 ± 0.25*

The percentages and cell numbers (x 10⁵ cells) in each compartment were determined by flow cytometry and expressed as the mean ± SEM. CD25 expression was determined based on CD4⁺ cells. The data presented here from 22 control mice (14 female and 8 male DBF₁ mice) and 21 mercury-exposed mice (12 female and 9 male DBF₁ mice) from three independent experiments. *p < 0.01 was the level of significance.

Figure 2.  Prenatal HgCl₂ exposure leads to increased proliferative responses in lymphocytes. Splenic lymphocytes (5 × 10⁵ cells) from mercury-exposed and unexposed (A) male and (B) female mice were stimulated with ConA (2 μg/ml), LPS (15 μg/ml) or idiopeptide (15 μg/ml) for 72 hr. Data were expressed as the proliferation index, computed as the ratio of the proliferative response of stimulated cells (measured as absorbance) to that of the unstimulated cells. Data were collected from 18 control mice (9 female and 9 male DBF₁ mice) and 23 mercury-exposed mice (12 female and 11 male DBF₁ mice) and shown as mean ± SD. The differences between mercury-exposed mice and sex-matched unexposed mice were considered significant when p < 0.05.

Figure 3.  The effects of prenatal mercury exposure on the production of T_H1 cytokines. In vitro production of (A) IFNγ and (B) IL-2 by lymphocytes (10⁶ cells/well) from mercury-exposed and unexposed mice that were stimulated with ConA (2 μg/ml) or idiopeptide (15 μg/ml) was measured after 24 hr (IFNγ) or 72 hr (IL-2). Data were collected from 18 control mice (9 female and 9 male DBF₁ mice) and 23 mercury-exposed mice (12 female and 11 male DBF₁ mice) and shown as mean ± SD. The differences between mercury-exposed mice and sex-matched unexposed mice were considered significant when p < 0.05.

Figure 4.  The effects of prenatal mercury exposure on the production of T_H2 cytokines. In vitro production of (A) IL-4 and (B) IL-10 by lymphocytes (10⁶ cells/well) from mercury-exposed and unexposed mice after stimulation with ConA (2 μg/ml) or idiopeptide (15 μg/ml) was measured after 72 hr. Data were collected from 18 control mice (9 female and 9 male DBF₁ mice) and 23 mercury-exposed mice (12 female and 11 male DBF₁ mice) and shown as mean ± SD. The differences between mercury- exposed mice and sex-matched unexposed mice were considered significant when p < 0.05.

Table 4.  The effect of prenatal mercuric chloride exposure on Id^LNF₁-reactivity in splenic CD4⁺ T-cells.

	CD4^+Id^LNF1^+
	Percentage of cells	Number of cells (×10^5)
Control Male	0.75 ± 0.10	2.19 ± 0.14
Mercury Male	1.02 ± 0.13	3.79 ± 0.22
Control female	0.76 ± 0.07	1.90 ± 0.95
Mercury Female	1.23 ± 0.11*	3.25 ± 0.41*

The percentages and cell numbers (x 10⁵ cells) in each compartment were determined by flow cytometry and expressed as mean ± SEM. The data presented here from 22 control mice (14 female and 8 male DBF₁ mice) and 21 mercury-exposed mice (12 female and 9 male DBF₁ mice) from three independent experiments. *p < 0.01 was the level of significance.