Osteoporosis, a disease that diminishes bone mineral density, affects approximately 10.2 million older adults. It increases the risk of fractures, leading to a $19 billion economic burden and a 36% increase in mortality rate. Therefore, fracture prevention in older adults is imperative. As the world shifts towards a western diet, understanding the impact of dietary fat on bone health is vital due to its potential to increase fracture risk. Our study aimed to explore the effects of unsaturated fatty acids on bone health at both cellular and macroscopic levels. We hypothesized that a higher ratio of ω6:ω3 fatty acids would negatively impact bone accrual, strength, nutrient trafficking, and osteoblast function. In the first aim, we used a Fat-1 transgenic mouse model, which endogenously converts ω-6 to ω-3 fatty acids. The data showed a sex-specific effect when reducing the ω-6 to ω-3 fatty acid ratio, with males showing a reduced fracture toughness, unlike females. Our findings did not support our hypothesis, but we did not account for potential changes in nutrient handling. In our second aim, we used the mckLPL transgenic mouse model, capable of altering fatty acid metabolism by increasing skeletal muscle fat oxidation. These mice were subjected to weight loss and weight regain study design with and without exercise after exposure to a ω6 diet. We observed increased lipid retention in bone during periods of acute overfeeding, but this was not attenuated in mckLPL mice. These results did not support our hypothesis but provided new insights into the role of bone in weight regain after weight loss. In our final aim, we examined the individual effects of ω3, ω6, and ω9 fatty acids on osteoblast function, which relies on ATP for successful bone formation. We found that ω-6 fatty acids had dual effects on osteoblast function while ω3 and ω-9 fatty acids did not significantly alter the cells. Overall, our data offers valuable insights into the complex relationship between fatty acids and bone health that will inform dietary recommendations for the prevention of osteoporosis