CD146+CD107ahigh define a highly secretory and immunosuppressive Mesenchymal Stem/Stromal Cells subset with enhanced therapeutic effects reversing synovitis and fat pad fibrosis through M1/M2 phenotypic shift

Abstract

Background & Aim Mesenchymal stem/stromal cells (MSC) are highly immunomodulatory and promising therapeutic candidates for numerous clinical indications. Translational advances of cell-based therapies require standardization, thus intrinsic heterogeneity of MSC may warrant concerns and yield inconsistent outcomes. Human bone marrow-derived MSC (BM-MSC) subpopulations have been discriminated based on their specific niche, however thorough functional analyses correlating potencies has yet to be determined. Methods, Results & Conclusion Methods Herein, we investigated the comprehensive functionality of BM-MSC (n=8) discerned by perivascular marker CD146, inadvertently average in crude BM-MSC outcomes. Initially, inflammatory challenge (i.e., priming) to crude BM-MSC, termed unfractionated (UNF), captured a baseline of signature responses including enhanced CD146+ expression. So, UNF preparation and immunoselected subpopulations based on CD146 positivity (CD146+ & CD146− cohorts) were assessed for phenotypic, transcriptional and secretome profiles, and in vitro/in vivo functional evaluations (immunopotency assay and reversing inflammatory and fibrotic events induced in the rat knee). Results Both primed UNF and CD146+ sorted cells enriched for a CD107ahigh, CXCR4high, and LepRhigh phenotype.Molecular evaluations evidenced CD146+ cells (not the CD146− cohort) exhibiting a highly immunomodulatory transcriptome and secretome, with enhanced secretory capacity (determined by CD107aHigh). This was functionally evident in robust immunosuppression of both stimulated PBMC and T lymphocytes while inducing significant frequencies of Regulatory T cells, providing evidence of CD146+CD107ahigh BM-MSC as the distinctively potent responders to injury and inflammation. This was supported in vivo using a rat model of knee synovium and infrapatellar fat pad (IFP) inflammation and fibrosis, efficiently reversed following CD146+ BM-MSC treatment (even at reduced cell doses – 1/10th) and largely failed by the CD146− counterpart. Mechanistically, CD146+ cells greatly promoted a pivotal M1-to-M2 macrophage phenotypic shift within synovium and IFP further supporting local immunomodulatory effects. Conclusion This study definitively supports that within crude BM-MSC, the CD146+CD107ahigh subpopulation, deemed ‘first responders’ facilitate high therapeutic efficacy. By selection of the CD146+ BM-MSC, our evidence implicates a highly translatable method that may reduce the need for high cell yields while improving outcomes.