Transcriptional profiling at single-cell resolution reveals diversity and regulatory networks of primary and secondary senescent cells

We have employed a single cell sequencing approach using 10x Genomics scRNAseq to study revealed distinct transcriptomic landscapes and senescence trajectories between primary and secondary senescence cells (SnCs). Primary SnCs exhibited enrichment in ECM remodeling and fibrosis-associated pathways, with markers such as COL1A1, ITGA2, FAS, HPGD, and PYGB, implicating them in tissue structural alterations and inflammation. In contrast, secondary SnCs upregulated inflammatory and metabolic reprogramming pathways, with markers including CXCL8, CSF2, BMP2, LAMA4, and PHKG1, indicating their role in senescence propagation via SASP. Pseudotime analysis delineated distinct senescence trajectories, where primary SnCs progressed toward fibrotic and ECM-driven states, while secondary SnCs followed inflammation- and metabolism-driven trajectory. Despite these differences, both SnC subtypes shared core regulatory networks governed by transcription factors (TFs) HMGA1, NFKB1, and JUNB, which regulate SASP activity, ECM remodeling, and senescence maintenance. This study provides a comprehensive single-cell transcriptional map of primary and secondary SnCs, uncovering key molecular differences, distinct senescence trajectories, and shared regulatory pathways. By identifying subtype-specific markers and TFs, our findings advance the understanding of senescence propagation and highlight potential targets for next-generation senotherapeutics. Instead of broadly eliminating all SnCs, future senotherapheutic strategies should selectively targeting primary SnC subtypes that drive secondary senescence, preventing its pathological propagation while preserving beneficial functions.