ITGBL1 and RPN1 Mark a Fibrotic NP Subpopulation with Coupled Integrin Signaling and N-Glycosylation Programs in IVDD.

BACKGROUND: FibroNPC is a degeneration associated subtype in intervertebral disc degeneration (IVDD), yet its markers and immune crosstalk remain poorly characterized. ITGBL1 and RPN1 recur as IVDD associated genes in bulk studies, but where they are expressed within the heterogeneous disc has not been resolved. METHODS: Three GEO datasets comprising 81 bulk profiles were integrated and screened by LASSO, SVM-RFE, and Random Forest, with external validation in an independent 15 sample cohort. Two human IVDD scRNA-seq cohorts were analyzed by subclustering, signature scoring, and within scRNA-seq per-subpopulation contribution analysis. CellChat, AUCell pathway scoring, and drug prediction were used as exploratory analyses; IL-1β stimulated HNPCs served as the in vitro model. RESULTS: Machine learning analysis prioritized ITGBL1 and RPN1 as consensus core genes (combined AUC = 0.925), with their differential expression further supported in an independent external cohort. Single cell mapping preferentially localized both genes to a FibroNPC subpopulation and associated this cell state with coupled integrin/focal adhesion and ER protein processing/N-glycosylation programs. Notably, although FibroNPCs comprised less than one-fifth of NP/AF cells, they contributed approximately 48% of total ITGBL1 expression and disproportionate shares of several canonical fibrotic transcripts. CellChat further inferred increased macrophage SPP1-integrin communication involving structural cell populations in IVDD. In IL-1β stimulated HNPCs, both ITGBL1 and RPN1 were increased at the mRNA and protein levels. CONCLUSION: ITGBL1 and RPN1 represent FibroNPC associated molecular markers linked to coupled integrin signaling and N-glycosylation programs in IVDD. The disproportionate contribution of FibroNPCs to ITGBL1 and fibrotic gene expression highlights the importance of cell state resolved localization beyond conventional bulk hub-gene identification. Further functional validation is required before therapeutic implications can be established.