Evaluation of etiology and effects of oxidative stress in long bone development in Ts65Dn and Dp(16)1Yey mouse models for Down syndrome
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Down syndrome (DS) is one of the most common genetic disorders in humans, resulting in phenotypes that affect the cardiac, nervous, and skeletal systems. Skeletal phenotypes that have been observed in DS individuals include shortened long bones, short stature, reduced bone mineral density, and increased risk of osteoporosis; however, the etiology is not well understood. Research suggests that some of these phenotypes, particularly shortened long bone length, originate during prenatal development. This study aimed to characterize long bone phenotypes during embryonic development and evaluated oxidative stress in the Dp(16)1Yey mouse model for DS. This work focused on embryonic day (E) 14.5 and E15.5, as this is the time during endochondral ossification at which cartilaginous anlagen is converted to mineralized bone. The evaluation of long bone phenotypes was conducted through staining of cartilage with Alcian blue and calcified bone with Alizarin red. Anlagen length was similar in Dp(16)1Yey and euploid samples at both stages but ossification presence was reduced in Dp(16)1Yey samples at E14.5 and slightly increased in Dp(16)1Yey samples at E15.5, although the results were not significant at the later stage. Because apoptosis of chondrocytes occurs during mineralization, this study also utilized terminal deoxynucleotyidyl transferase dUTP nick end labling (TUNEL) to examine patterns of apoptosis and the state of chondrocyte terminal differentiation. This work was supplemented with an evaluation of oxidative stress as a factor that may contribute to skeletal phenotypes. Analysis of oxidative stress included quantitative real-time PCR to evaluate transcript abundance of oxidative stress response genes and quantification of copper/zinc superoxide dismutase levels, which is involved with antioxidant defense, through an enzyme linked immunosorbent assay. qRT-PCR results suggest the presence of oxidative stress in Dp(16)1Yey anlagen. Taken together, these findings suggest a delay in ossification of Dp(16)1Yey samples at E14.5 that is recovered and possibly reversed by E15.5, indicating altered rates of terminal differentiation of chondrocytes in Dp(16)1Yey long bones. Oxidative stress at these stages may be influencing this process. These results, along with future studies, may help to demonstrate embryonic origins of DS skeletal phenotypes in Dp(16)1Yey mice and contribute to the development of therapeutic treatments for DS individuals.
Franklin and Marshall College Archives, Undergraduate Honors Thesis 2018
- F&M Theses Collection