2003 – PHYSICALLY INTERACTING CELL SEQUENCING IN BONE MARROW FIBROSIS REVEALS DISTINCT CELLULAR NICHES IN FIBROSIS

• Published in: Experimental hematology 2024-08, Vol.137, p.104560, Article 104560
• Main Authors: Gleitz, Helene, Nagai, James, Ruiz, Mayra, Fuchs, Stijn, Snoeren, Inge, Boeree, Aurélie, Cesaro, Giulia, Kramann, Rafael, Costa, Ivan, Schneider, Rebekka
• Format: Article
• Language: English
• ISSN: 0301-472X
• DOI: 10.1016/j.exphem.2024.104560

Recent studies highlight cellular crosstalk between fibrosis-inducing hematopoietic cells and fibrosis-driving stromal cells in bone marrow (BM) niche transformation in myeloproliferative neoplasms (MPNs). Absence of CXCL4 (PF4) reduces stromal cell activation, BM fibrosis and profibrotic pathways in megakaryocytes (MKs). Understanding this crosstalk is crucial for therapeutic targeting, but challenging as signaling pathways dependent on direct physical interactions are lost in current single-cell sequencing protocols. We aimed to dissect physical interactions between PF4-expressing cells (megakaryocytes, macrophages, monocytes) and Gli1+ stromal cells. Genetic fate tracing and BM transplantation from Pf4-ZsGreen mice into Gli1CreERt2;tdTomato mice tracked cell fate post-transduction with TPO vector inducing BM fibrosis. As mice showed fibrosis symptoms, Gli1+ cells and Pf4+ cells were sorted individually (singlets) and as multiplets (tdTomato+ ZsGreen+). We reveal an unprecedented representation of monocytes, macrophages, CAR stromal cells and endothelial cells in BM fibrosis with 27,000 singlets and 35,927 physically interacting multiplets. Computational analysis of singlets and multiplets identified changes in physically interacting cells and underlying cellular communication networks in disease. Computational analysis and imaging reveal various interacting niches that adapt in disease, and a macrophage subtype that acts as an "interaction hub," upregulating Spp1 expression in macrophages and osteoCAR cells. By directly targeting this interaction using an Spp1 genetic knockout in HSCs, we show an amelioration in BM fibrosis and a reduced anemic phenotype, hallmarks of advanced MPN. This novel approach allows us to decipher intricate cellular interactions in BM fibrosis pathogenesis and identify novel therapeutic targets that directly impact BM fibrosis.