Molecular mechanisms of space related bone loss.

摘要

Mechanical loading is a major player in the maintenance of a normal bone phenotype. Increasing load or force on bone promotes bone formation while decreasing load promotes bone loss. The molecular events mediating responses to alterations in skeletal loading are largely unknown. My thesis focuses on determining the molecular effects of altered mechanical load on bone forming osteoblasts using a system of decreased load.; The NASA designed rotating wall vessel (RWV) allows for the culture of cells in solid phase rotation around a horizontal axis such that gravity vectors are randomized to near zero thereby modeling microgravity. Osteoblasts exposed to 24 hours of RWV conditions exhibit suppressed expression of markers of osteoblast differentiation, namely alkaline phosphatase and osteocalcin. Furthermore, transcription factors that regulate the expression of these genes and progression to a fully differentiated osteoblast are also suppressed in the RWV. Specifically, c-fos and c-jun expression, members of the AP-1 family, are suppressed after 24 hours of RWV culture. Consistent with this finding AP-1 DNA binding and transactivation are suppressed. The osteoblast transcription factor runx2 displays similar effects to AP-1 in modeled microgravity conditions. After 24 hours of RWV culture, runx2 mRNA levels are decreased along with runx2 DNA binding and runx2 promoter activation. This demonstrates that conditions of decreased loading may be involved in suppressed osteoblast phenotype through alterations in transcription factor activities.; While the RWV models microgravity, there is also a hypoxic component associated with the culture of differentiating osteoblasts. To uncouple the effects of hypoxia from modeled microgravity, external oxygen was supplied to RWV cultured cells to normoxia. Under normoxia, the suppression of c-fos, c-jun, and runx2 expression does not occur suggesting that the hypoxic component of the RWV and not modeled microgravity mediates the initial suppression of these genes we observed. Interestingly, skeletal unloading induces a hypoxic microenvironment to which bone cells are subjected. My findings suggest that bone loss associated with unloading, as seen during space flight, may be directly due to hypoxia.