Effect of High Glucose and Carboxymethyl-Lysine on Osteocyte Gene Expression

Abstract

Diabetes mellitus (DM) is associated with increased bone fragility despite normal or elevated bone mineral density, partially due to the accumulation of advanced glycation end products (AGEs) in bone tissue. AGEs, such as carboxymethyl lysine (CML), impair osteocyte function by activating the receptor for advanced glycation end products (RAGE), triggering oxidative stress and inflammatory responses. This study aimed to investigate the effects of high glucose (HG) and CML on bone remodeling, glycation, inflammatory markers, and cellular functions in osteocytes. Using the murine osteocyte cell line OCY454-12H, we treated cells with HG (30 mM glucose) or 3 µM CML to simulate diabetic conditions. We assessed the expression of bone remodeling markers (SOST, RANKL, OPG, CTsK), glycation markers (RAGE, AGER1), inflammatory cytokines (IL-6, TNF- α ), and cellular functions, including proliferation, viability, and apoptosis, using quantitative PCR and functional assays. HG treatment resulted in a 10-fold increase in SOST expression (9.3 vs. 0.9, p ≤ 0.0001) and a 2.4-fold increase in RANKL expression (2.75 vs. 1.15, p ≤ 0.0001), with a concurrent 2-fold increase in OPG (2.60 vs. 1.04, p ≤ 0.0001). The RANKL/OPG ratio remained unchanged (p = 0.15). HG also significantly increased RAGE expression by 3.67-fold (4.20 vs. 1.15, p ≤ 0.0001) and AGER1 by 1.65-fold (1.94 vs. 1.15, p ≤ 0.0001), along with a 2.02-fold increase in IL-6 (2.32 vs. 1.12, p ≤ 0.001) and a 7.35-fold increase in TNF- α (7.04 vs. 1.04, p ≤ 0.0001). Cell viability and proliferation were significantly higher under HG, accompanied by increased caspase-3 activity, indicating enhanced apoptosis. In contrast, CML exposure significantly upregulated RAGE (3.18 vs. 1.15, p ≤ 0.0001) and AGER1 (2.10 vs. 1.14, p = 0.028) but had no significant effects on bone remodeling markers, inflammatory cytokines, or cellular functions at physiological concentrations. Our findings demonstrate that HG disrupts osteocyte function by altering bone remodeling, glycation, and inflammatory pathways, while CML at physiological levels selectively activates glycation markers without inducing broader cellular dysfunction. These results underscore the role of the AGE-RAGE axis in diabetic bone fragility and highlight the need for future in vivo studies to explore therapeutic strategies targeting AGE accumulation and RAGE signaling in bone.