Simulated microgravity models sex differences in knee osteoarthritis through a CD36-regulated mechano-metabolic circuit

Sex differences in knee osteoarthritis (KOA) are well documented, but the molecular drivers within meniscal tissue remain unclear. We studied whether simulated microgravity (SMG) models sex‑dependent meniscal degeneration and whether the lipid scavenger receptor CD36 mediates these effects. Human menisci from total knee arthroplasty (TKA) donors were analyzed (histology n = 5 females/5 males; flow cytometry and RT‑qPCR n = 7 females/7 males). In mice, we assessed spontaneous anterior‑horn mineralization by micro‑CT at 4, 12, and 24 weeks in wild‑type (WT) and CD36‑deficient (Cd36−/−) animals. Four‑week menisci were then encapsulated in 1% agarose and cultured for 3 weeks under static or SMG conditions with serial micro‑CT (days 0, 7, 14, 21), histology, bulk RNA‑seq, and DIA shotgun proteomics. Female TKA menisci showed near‑complete glycosaminoglycan loss, disorganized type I collagen, and elevated hypertrophic markers (COL10A1, BMP2, IHH), with a trend toward more CD36 + cells. In vivo, a significant sex and genotype interaction emerged at 12 weeks in mouse model: Cd36−/− females displayed ~ 15% less mineralization than WT. Ex vivo, SMG increased mineralization in WT females by ~ 25% at day 7 and ~ 30% at day 21 compared to static, but had minimal effect in males; Cd36−/− females exhibited ~ 20–30% lower SMG‑induced mineralization than WT and preserved Safranin‑O and type II collagen. Transcriptomics under SMG revealed strong sex divergence: females showed enrichment of cell–matrix/mechanosignaling pathways, including ECM–receptor/focal‑adhesion and Ca/PI3K‑linked signaling, whereas males showed immune/osteoclast signatures. Proteomics paralleled these patterns: WT females under SMG had marked decreases in Col2a1, Col11a1, and Acan with increased oxidative‑phosphorylation proteins; Cd36−/− females showed reduced abundance of mitochondrial respiratory proteins and vesicle/mineralization‑associated factors, consistent with reduced mineralization and preserved ECM. SMG provides a controllable platform that recapitulates sex‑dependent, KOA‑like tissue degeneration. The data support a CD36-dependent mechanometabolic axis in females in which mechanical unloading disrupts ECM integrity, activates CD36-linked lipid handling with Ca/PI3K/AMPK signaling, and drives higher mitochondrial respiration and oxidative stress, culminating in ECM breakdown and mineralization, positioning CD36 as a promising sex-stratified therapeutic target in KOA.