Transfer learning of an in vivo-derived senescence signature identifies conserved and tissue-specific senescence across species and diverse pathologies

Senescent cells (SnCs) contribute to normal tissue development and repair but accumulate with aging where they are implicated in a number of pathologies and diseases. Despite their pathological role and therapeutic interest, SnC phenotype and function in vivo remains unclear due to the challenges in identifying and isolating these rare cells. Here, we developed an in vivo -derived senescence gene expression signature using a model of the foreign body response (FBR) fibrosis in a p16Ink4a- reporter mouse, a cell cycle inhibitor commonly used to identify SnCs. We identified stromal cells (CD45-CD31- CD29+) as the primary p16Ink4a expressing cell type in the FBR and collected the cells to produce a SnC transcriptomic signature with bulk RNA sequencing. To computationally identify SnCs in bulk and single-cell data sets across species and tissues, we used this signature with transfer learning to generate a SnC signature score (SenSig). We found senescent pericyte and cartilage-like fibroblasts in newly collected single cell RNAseq (scRNASeq) data sets of murine and human FBR suggesting populations associated with angiogenesis and secretion of fibrotic extracellular matrix, respectively. Application of the senescence signature to human scRNAseq data sets from idiopathic pulmonary fibrosis (IPF) and the basal cell carcinoma microenvironment identified both conserved and tissue-specific SnC phenotypes, including epithelial-derived basaloid and endothelial cells. In a wound healing model, ligand-receptor signaling prediction identified putative interactions between SnC SASP and myeloid cells that were validated by immunofluorescent staining and in vitro coculture of SnCs and macrophages. Collectively, we have found that our SenSig transfer learning strategy from an in vivo signature outperforms in vitro -derived signatures and identifies conserved and tissue-specific SnCs and their SASP, independent of p16Ink4a expression, and may be broadly applied to elucidate SnC identity and function in vivo . ### Competing Interest Statement J.H.E. holds equity in Unity Biotechnology and Aegeria Soft Tissue and is an advisor for Tessera Therapeutics, HapInScience, and Font Bio. D.M.P. is consultant at Aduro Biotech, Amgen, Astra Zeneca, Bayer, Compugen, DNAtrix, Dynavax Technologies Corporation, Ervaxx, FLX Bio, Immunomic, Janssen, Merck, and Rock Springs Capital. D.M.P. holds equity in Aduro Biotech, DNAtrix, Ervaxx, Five Prime therapeutics, Immunomic, Potenza, Trieza Therapeutics. D.M.P. is a member of the scientific advisory board for Bristol Myers Squibb, Camden Nexus II, Five Prime Therapeutics, and WindMil. D.M.P. is a member of board of directors in Dracen Pharmaceuticals. C.C. is the founder and owner of C M Cherry Consulting, LLC. E.J.F. is a member of the scientific advisory board for Resistance Bio and is a consultant for Merck and Mestag Therapeutics. J.M.v.D. is a co-founder of and holds equity in Unity Biotechnology and Cavalry Biosciences. D.J.B. is a shareholder and co-inventor on patent applications licensed to or filed by Unity Biotechnology, a company developing senolytic medicines, including small molecules that selectively eliminate senescent cells. Research in his lab has been reviewed by the Mayo Clinic Conflict of Interest Review Board and is being conducted in compliance with Mayo Clinic Conflict of Interest policies.