Type 2 diabetes mellitus (T2DM) increases the susceptibility of bone fragility. The underlying mechanisms have, however, remained largely unknown. MicroRNAs (miRNAs) are short single-stranded non-coding RNA molecules with utility as biomarkers due to their easy accessibility and stability in bodily fluids. Here, we aimed to use an unbiased approach to identify miRNAs dysregulated in a polygenic mouse model of T2DM. Genome-wide analysis of miRNAs in serum, BM, and bone from the polygenic TallyHo/JngJ (TH) mice, which recapitulate T2DM in humans, was performed. This analysis was compared to the recommended control Swiss Webster Random/Jackson (SWR/J) and a strain-matched non-diabetic control (TH-ND). When comparing TH mice with TH-ND using an adjusted p-value false discovery rate (FDR) cut-off of 0.2 to identify differentially expressed miRNAs, mmu-miR-466i-5p and mmu-miR-1195 were found to be up-regulated in both serum and in BM. Dysregulated miRNAs were not found in bone tissue. When comparing TH-ND mice with SWR/J using the same FDR cut-off, mmu-miR-351-5p, and mmu-miR-322-3p were upregulated in both BM and serum, while mmu-miR-449a-5p and mmu-miR-6240 were downregulated in BM and serum. Dysregulated miRNAs in BM or cortical bone compared to serum between TH-ND mice and SWR/J were investigated for their cell-type enrichment to identify putative donor cells and their gene target networks. Gene target network analysis revealed genes involved in diabetes-related signaling pathways as well as in diabetic bone disease. Cell-type enrichment analysis identified hsa-miR-449a enriched in immune cells, hsa-miR-592 in hepatocytes and endothelial cells, while hsa-miR-424-3p, hsa-miR-1-3p, and hsa-miR-196b-5p were enriched in mesenchymal stem cells and their derived tissues. In conclusion, our comparative miRNA profiling sheds light on differential expression patterns between SWR/J and both subgroups of TH. No differences were observed between TH and TH-ND, suggesting the genetic background of SWR/J may be responsible for the change of dysregulated miRNA.