Recurrent cholesterol-enriched cartilage signature and SREBF1-sensitive lipid and catabolic responses in osteoarthritis

Objective The aim in this study was to identify a sterol-associated transcriptional program recurrent across human osteoarthritis (OA) cartilage datasets and test whether SREBF1 influences lipid metabolism and catabolic responses in chondrocytes. Methods We used GSE220243 for donor-level, state-resolved discovery, GSE169454 for independent donor-level testing, and GSE114007 for platform-adjusted analysis of whole-cartilage RNA-seq profiles. We prioritized the cholesterol-homeostasis-associated module PM214 on biological grounds and evaluated its 27-gene signature across cohorts. SREBF1 activity inference was accompanied by a sensitivity analysis excluding shared regulon genes. Two independent SREBF1 siRNAs were tested in C28/I2 cells by RT-qPCR (six independent experiments) and immunoblotting (four experiments). Further experiments measured cellular cholesterol, triglycerides, MMP13 secretion, and the effects of genetic re-expression under interleukin-1β (IL-1β) stimulation (six independent batches per module). Results The Hallmark cholesterol-homeostasis component of PM214 was enriched in hypertrophic/ossifying chondrocytes (NES 2.124; FDR 3.74 × 10 −4 ; 19 leading-edge genes). The 27-gene PM214 score was higher in OA-associated profiles in GSE169454 (exact P = 0.0286) and GSE114007 (platform-adjusted P = 8.00 × 10 −4 ). Although SREBF1 was prioritized by transcription-factor activity analysis, its correlation with PM214 declined from 0.797 to 0.224 after eight shared genes were excluded (matched-null P = 0.582). Both siRNAs reduced four prespecified sterol-related transcripts, mature SREBP1 and HMGCR protein, cellular total cholesterol and triglycerides, and IL-1β-stimulated MMP13 secretion. Re-expression of siRNA-resistant SREBF1 restored key cholesterol, triglyceride, and MMP13 responses. Conclusion PM214 is a recurrent OA-associated cartilage expression signature with a prominent cholesterol-homeostasis component. SREBF1 perturbation and rescue implicate this factor in lipid and catabolic responses in C28/I2 cells, without establishing direct regulation of the complete PM214 program or effects in primary human cartilage.