Abstract
Purpose
Developmental exposure to estrogens has been shown to affect a number of organ systems, including long and short bones. Epigenetic effects of DES exposure have been shown to affect the third generation of progeny. Furthermore, recent studies have shown that environmental exposure to estrogen-like compounds is much higher than originally anticipated. This study aims to discover the effect of in utero exposure to a well-known estrogen agonist, diethylstilbestrol (DES), on lumbar bone, intervertebral disc (IVD), and articular cartilage. Femoral bone was studied to determine the specificity of the effect.
Method
C57bl/6n pregnant mice were dosed orally with vehicle (peanut oil) or 0.1, 1.0 and 10 g/kg/day of DES on gestational days 11–14. Male and female pups were allowed to mature without further treatment until 3 months of age, at which point they were divided into swim and sedentary groups. After sacrifice, bone mineral density (BMD), bone mineral content (BMC), bone area (BA), and trabecular bone area (TBA) of the lumbar vertebrae and femur were measured using a PIXImus Bone Densitometer System (GE Medical Systems). Glycosaminoglycan (GAG) content (proteoglycan) was measured by the DMMB assay. Histological analysis of proteoglycan was performed with Safranin O staining. Intervertebral disc height was measured using NDP software (Leeds, UK). Statistical analysis was performed using analysis of variance (ANOVA) followed by Fisher's Protected Least Significant Difference (PLSD). A p-value of < 0.05 was considered statistically significant.
Results
At all the doses studied, DES had no significant effect on lumbar and femoral BMD in both males and females. The lumbar BMC, however, was significantly increased in female swims at both the highest and lowest dose of DES, while the femoral BMC was only increased at the highest. The males, on the other hand, showed a decreased BMC at the highest dose of DES for both lumbar and femoral bone. Female swim group had an increased BA at the highest dose of DES while the male swims showed a decreased BA for femoral bone. The TBA showed a similar pattern. GAG analysis of lumbar IVDs showed a decrease at the lowest doses but a significant increase at the highest doses for both swim and sedentary groups. Histology showed morphological changes of the IVD and articular cartilage for all doses of DES.
Conclusion
The effect of in utero DES exposure is more important on lumbar than on femoral bone. The effect was mainly observed at high dose of DES, except for BA that is also affected by low dose DES in lumbar bones. Results suggest that environmental estrogen contaminants might impact developmental lumbar bone growth and mineralization in mice. Further studies measuring the impact of environmental estrogen mimics, such as bisphenol A, are then warranted.
