Impacts of Bisphenol A and Ethinyl Estradiol on Male and Female CD-1 Mouse Spleen

Abstract

The endocrine disruptor bisphenol A (BPA) and the pharmaceutical 17α-ethinyl estradiol (EE) are synthetic chemicals with estrogen-like activities. Despite ubiquitous human exposure to BPA, and the wide-spread clinical use of EE as oral contraceptive adjuvant, the impact of these estrogenic endocrine disrupting chemicals (EDCs) on the immune system is unclear. Here we report results of in vivo dose response studies that analyzed the histology and microstructural changes in the spleen of adult male and female CD-1 mice exposed to 4 to 40,000 μg/kg/day BPA or 0.02 to 2 μg/kg/day EE from conception until 12-14 weeks of age. Results of that analysis indicate that both BPA and EE have dose- and sex-specific impacts on the cellular and microanatomical structures of the spleens that reveal minor alterations in immunomodulatory and hematopoietic functions. These findings support previous studies demonstrating the murine immune system as a sensitive target for estrogens, and that oral exposures to BPA and EE can have estrogen-like immunomodulatory affects in both sexes.

Figure 1

Effects of exposure on spleen weight. Spleen weight of males and females exposed to increasing doses of BPA (A) or EE (B). *Indicates significantly different from same sex control p < 0.05 by Dunnett’s multiple comparisons test.

Figure 2

Representative photomicrographs of the splenic microanatomical structures in control and 17α-ethinyl estradiol exposed adult male and female CD-1 mice. Shown are representative H&E stained sections of splenic tissue from (A) control male and (B) control female mice illustrating the normal micro-architecture of the white pulp (arrow) and red pulp (asterisk). Note that control tissue displays abundant white pulp components. (C) Representative images of males exposed to dietary 17α-ethinyl estradiol (0.01 ppm EE). Decreased periarteriolar lymphoid sheath (PALS) numbers, size and cellularity (arrow), together with paucicellular primary lymphoid follicles (arrowhead) are evident. (D) Shown is an image for a representative female from the 0.01 ppm EE exposure group showing decreased PALS numbers and cellularity (arrow) with discrete primary lymphoid follicles and mildly decreased numbers of follicular lymphocytes (arrowhead). (E) Representative image of an EE-exposed male from the 0.01 ppm exposure group which demonstrates increased amounts of golden-yellow pigment (hemosiderin) in the red pulp that was often contained within macrophages (arrows and inset). (F) Representative image from a 0.0001 ppm EE exposed female demonstrating the observed phenotype of diffuse expansion of the red pulp by extramedullary hematopoiesis (EMH). The EMH was characterized by the presence of myeloid, megakaryocytic precursors, and a morphological predominance of erythroid precursors (inset). Scale bars indicate 100 µm.

Figure 3

Representative photomicrographs of splenic microanatomical structures in EE and BPA-exposed adult male and female CD-1 mice. Shown are representative H&E stained sections of splenic tissue from (A) a male mouse exposed to BPA (0.03 ppm), and (B) a female mouse exposed to 3 ppm BPA. These are illustrative of the representative phenotypes in males and females that was characterized by decreased PALS (arrowhead) numbers, size, and lymphocyte cellularity (arrow). These effects were typically more pronounced and diffuse in male versus females. (C) High magnification image of a male spleen representative of the 0.03 ppm BPA exposure group and demonstrating decreased PALS (arrowhead) size, numbers and lymphocyte cellularity (arrow), along with decreased primary lymphoid follicles cellularity (arrow). (D) High magnification image of a representative female spleen from the 3 ppm BPA exposure group characterized by decreased PALS numbers and cellularity with decreased lymphoid follicle cellularity. In addition to the white pulp changes, BPA-exposed females also had increased hematopoietic precursor populations (EMH) within the red pulp, which were predominantly of erythroid lineage (asterisks). Shown is a representative image of a section from a 0.0001 ppm EE-exposed male (E) and female from the 0.03 BPA group (F) showing prominence of marginal zones, a phenotype that was observed in primarily males from EE exposure groups and female from both BPA- and EE-groups. Scale bars indicate 100 µm.