Sodium Methyldithiocarbamate Causes Thymic Atrophy by an Indirect Mechanism of Corticosterone Up-Regulation

Figures & data

TABLE 1 Thymocyte and splenocyte subpopulation percentages and cell counts after SMD exposure

	CD4^−CD8^−	CD4^+CD8^−	CD4^−CD8^+	CD4^+CD8^+
a. Thymus
Naïve	1.7 ± 0.3 × 10^6.	8.6 ± 0.9 × 10^6.	3.3 ± 0.2 × 10^6.	65.2 ± 4.0 × 10^6.
	(2.2 ± 0.8.%)	(10.8 ± 0.5.%)	(4.1 ± 0.2.%)	(82.4 ± 0.7.%)
SMD 1.×	1.6 ± 0.1 × 10^6.	9.0 ± 0.3 × 10^6.	3.7 ± 0.2 × 10^6.	32.8 ± 2.2 × 10^6.
	(3.5 ± 1.3.%)	(18.6 ± 0.7.%)	(7.6 ± 0.6.%)	(68.2 ± 1.6.%)
SMD 3.×	1.8 ± 0.5 × 10^6.	8.7 ± 0.3 × 10^6.	2.7 ± 0.5 × 10^6.	12.6 ± 2.3 × 10^6.
	(6.8 ± 2.1.%)	(33.4 ± 4.1.%)	(10.3 ± 1.2.%)	(48.1 ± 6.5.%)
b. Spleen
Naïve	46.4 ± 3.3 × 10^6.	12.2 ± 0.7 × 10^6.	6.5 ± 0.5 × 10^6.	0.02 ± 0.007 × 10^6.
	(72.0 ± 0.7.%)	(18.0 ± 0.6.%)	(10.0 ± 0.2.%)	(< 0.1.%)
SMD 1.×	44.5 ± 5.1 × 10^6.	8.7 ± 1.0 × 10^6.	4.9 ± 0.5 × 10^6.	0.02 ± 0.002 × 10^6.
	(76.2 ± 1.0.%)	(15.2 ± 0.7.%)	(8.6 ± 0.3.%)	(< 0.1.%)
SMD 3.×	38.7 ± 0.2 × 10^6.	7.7 ± 0.1 × 10^6.	4.8 ± 0.2 × 10^6.	0.01 ± 0.007 × 10^6.
	(75.0 ± 0.7.%)	(15.4 ± 0.3.%)	(9.6 ± 0.4.%)	(< 0.1.%)

Mice were exposed to SMD (300 mg/kg) one time (1×) or three times (3×). Thymocytes (a) and splenocytes (b) were collected 72 hours after the 1 single dosage of SMD (1×) or 24 hours after the last (3×) SMD administration. Percent of each subpopulation is listed below in parenthesis. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 1 Dose-response effect of SMD on thymic and splenic cellularity. Mice were administered SMD orally at 9:00 PM daily for 3 days. On day 4 thymi (a) and spleens (b) were collected and cellularity was determined. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 2 SMD-increased serum corticosterone in mice. Mice were administered SMD (300 mg/kg) orally at 9:00 PM. After 1, 2, 4, 6, or 8 hours, trunk blood was collected and serum was isolated for corticosterone analysis by RIA. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 3 SMD did not cause thymic atrophy in adrenalectomized mice. Adrenalectomized mice were administered SMD (200 mg/kg) orally at 9:00 PM on day 1 only. On day 3, thymi and spleens were removed and analyzed for alterations in cellularity. Corticosterone levels were analyzed for all samples. Two samples (each from a different group) were omitted due to the presence of serum corticosterone, indicative of adrenal regeneration. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 4 Aminoglutethimide blocked serum corticosterone for up to 8 hours in vivo. Mice were administered aminoglutethimide for 8, 4, and 1 hour. One hour before collection of serum, all mice (except naïve) were restrained to induce a stress response. Trunk blood was collected at 9:00 AM and serum was isolated for analysis of corticosterone by RIA. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 5 SMD did not cause thymic atrophy in combination with aminoglutethimide. Mice were pretreated with aminoglutethimide 1 hour before SMD administration. SMD was administered (200 mg/kg) daily for 3 days. On day 4, thymi (a) and spleens (b) were collected and analyzed for alterations in cellularity. N= 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

TABLE 2 Aminoglutethimide blocks alterations in thymic subpopulations caused by SMD

	CD4^−CD8^−	CD4^+CD8^−	CD4^−CD8^+	CD4^+CD8^+
a. Thymus
Naïve	5.0 ± 0.4	13.5 ± 0.4	6.0 ± 0.6	75.5 ± 1.1
SMD	10.29 ± 1.4	38.3 ± 6.4	9.7 ± 1.9	41.7 ± 9.7
Aminoglu	3.9 ± 0.2	12.8 ± 0.2	4.1 ± 0.2	79.3 ± 0.2
SMD + Aminoglu	3.5 ± 0.2	13.4 ± 0.7	3.5 ± 0.2	79.6 ± 0.7
b. Spleen
Naïve	62.9 ± 1.5	24.4 ± 1.1	11.8 ± 0.6	< 0.1
SMD	56.8 ± 2.8	27.2 ± 1.5	15.4 ± 1.2	< 0.1
Aminoglu	60.9 ± 2.1	26.7 ± 1.5	11.5 ± 0.8	< 0.1
SMD + Aminoglu	67.7 ± 2.4	22.3 ± 1.6	9.2 ± 0.8	< 0.1

Mice were exposed to SMD (200 mg/kg) daily for 3 days with or without aminoglutethimide. Thymocyte (a) and splenocyte (b) subpopulations were collected 24 hours after last SMD dosage. Percent of each subpopulation is listed. N = 5 mice per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 6 Aminoglutethimide blocked serum corticosterone up-regulation by SMD. Mice were pretreated with aminoglutethimide for 1 hour before SMD administration. SMD was administered (200 mg/kg) by oral gavage. After 1 hour, trunk blood was collected and serum was analyzed by RIA for serum corticosterone. N = 5 per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).

FIG. 7 SMD intoxication at a high and low dosage in combination with a psychogenic/neurogenic stressor was not synergistic. Mice were dosed with SMD (200 or 50 mg/kg) at 9:00 PM for 3 consecutive days. Immediately after SMD dosage, mice were restrained for 2 hours in their home cages. On day 4, thymi and spleens were removed and analyzed for alterations in cellularity. N = 5 per group. Asterisks (*) indicate significant difference from naïve control (p < 0.05).