Canagliflozin-induced sex-specific adaptive metabolism in bone

Sodium glucose transporter-2 inhibitors (SGLT2i) were designed to increase the excretion of glucose through urine and lower blood glucose levels independent of insulin. Beyond their primary functions, SGLT2i have been observed to induce a wide range of metabolic effects. To understand how SGLT2i-mediated metabolic changes affect skeletal metabolism, six months old genetically diverse UM-HET3 mice were treated with canagliflozin (CANA), a SGLT2i, for 1, 3, and 6 months. The comprehensive analysis included assessments of serum hormones, morphological and mechanical properties of long bones and vertebrae, and the bone tissue metabolome. The findings demonstrated significant sex-specific metabolic adaptations to CANA treatment, evident as early as one-month post-treatment. Male mice treated with CANA exhibited notable reductions in body weight, serum leptin, and insulin levels, alongside decreases in proinflammatory markers TNFa and MCP-1. These changes were accompanied by decreased serum levels of bone remodeling markers P1NP and CTX, and by reduced periosteal bone remodeling as determined by histomorphometry. The bone tissue metabolome analysis further underscored distinct sex-specific metabolic responses and adaptations to CANA treatment. Specifically, CANA-treated female mice demonstrated enhanced adenine and adenosine salvage pathways and purine metabolism relative to female controls. In contrast, CANA-treated male mice exhibited increased activity in the FXR/RXR pathway and the GABAergic receptor signaling pathways compared to their control counterparts. Notably, the most significant difference in the metabolome between CANA-treated males and females was observed in the Branched-Chain Amino Acid (BCAA) Catabolism pathway. This study highlights the novel finding of sex-specific metabolic and skeletal adaptations to SGLT2i treatment, providing new insights into the complex interplay between glucose metabolism and bone health, while emphasizing the need for tailored therapeutic strategies that consider sex-specific responses to SGLT2i treatment. Overall design: To test the effects of CANA treatment on the potential of bone MSCs to differentiate to osteoblasts, bone marrow was isolated from mice treated with control or CANA-containing diet for three months. Bone marrow MSCs cultured for 7-days in ?-modified minimum essential medium (?MEM, CAT#) containing 10% fetal calf serum (FBS), penicillin/sterptomycin and fungizone. At day 8 cells were cultured in osteoblast induction media containing: ?MEM, 10%FBS, penicillin/streptomycin, fungizone, dexamethasone (Sigma Cat # D8893), ß-glycerol phosphate (Sigma Cat # 50020), and ascorbic acid (Sigma cat# A4403-100MG). Cells from each mouse were cultured for several assays: 1) RNAseq analysis following 10 days in culture, 2) alkaline phosphatase staining (Sigma cat#) following 14 days in culture, 3) alizarin red staining (Sigma cat#), following 21 days in culture 4) VonKosa staining (Sigma cat#) following 21 days in culture.