Altered tibial microRNA profiles and local bone remodeling markers in Zucker rat models of obesity and type 2 diabetes

Obesity and type 2 diabetes mellitus are associated with adverse skeletal outcomes, including an increased risk of fragility fractures, but the molecular changes underlying bone dysfunction remains incompletely understood. MicroRNAs (miRNAs) are post-transcriptional regulators with important roles in bone development and remodeling. We characterized tibial bone miRNA profiles and local bone remodeling markers in Lean, obese nondiabetic Fatty, and obese diabetic Fatty Zucker rats to identify molecular patterns shared by the two obese phenotypes and those associated with the diabetic phenotype. Tibial bone was analyzed by small RNA sequencing, computational target prediction, functional enrichment analysis, and quantitative real-time PCR of selected miRNAs. Osteocalcin and C-terminal telopeptide of type I collagen were measured as local markers related to bone formation and resorption, respectively. Sequencing detected 850 miRNAs, including 499 annotated and 351 candidate novel miRNAs. An exploratory analysis identified 102 miRNAs that met q < 0.05 in each of the three pairwise comparisons, of which 14 also met |log 2 FC| > 1 in every comparison. Five miRNAs displayed consistent ordered group-wise expression: rno-let-7c-2-3p and novel-rno-miR-446-5p increased across the Lean, Fatty, and Diabetic Fatty groups, whereas rno-let-7a-1-3p, novel-rno-miR-39-3p, and novel-rno-miR-57-3p decreased. Notably, novel-rno-miR-446-5p also met q < 0.05 and |log 2 FC| > 1 in both Lean-referenced comparisons, identifying it as the principal novel candidate. Sequencing and quantitative real-time PCR showed directional agreement in eight of ten evaluated contrasts, although only four contrasts reached statistical significance. Predicted targets were enriched in signaling pathways relevant to skeletal metabolism, including Wnt, PI3K–Akt, MAPK, mTOR, and Hippo, but these findings remain hypothesis generating. Fatty and Diabetic Fatty rats had lower tibial osteocalcin concentrations, while Diabetic Fatty rats also had higher C-terminal telopeptide of type I collagen concentrations. Because the study was cross-sectional and both obese groups shared the homozygous fa/fa mutation in Lepr , the observed differences cannot be attributed exclusively to obesity or diabetes or interpreted as disease progression. Within these constraints, the findings expand the miRNA landscape of bone and identify novel-rno-miR-446-5p as a candidate for independent validation and functional investigation in obesity- and diabetes-associated skeletal alterations.