Abstract
Background: To investigate the in vitro and in vivo anti-inflammatory/anti-fibrotic capacity of IFP-MSC manufactured as 3D spheroids. According to our hypothesis, IFP-MSC do not require prior cell priming to acquire a robust immunomodulatory phenotype in vitro in order to efficiently reverse synovitis and IFP fibrosis and secondarily delay articular cartilage damage in vivo. Methods: Human IFP-MSC immunophenotype, tri-potentiality, and transcriptional profiles were assessed in 3D settings. Multiplex secretomes were assessed in IFP-MSC spheroids [Crude (non-immunoselected), CD146+ or CD146- immunoselected cells] and compared with 2D cultures with and without prior inflammatory/fibrotic cell priming. Functionally, immunopotency limiting human PBMCs proliferation and effect on stimulated synoviocytes with inflammation and fibrotic cues. Finally, spheroids were tested in vivo in a rat model of acute synovitis/fat pad fibrosis. Results: Spheroids enhanced IFP-MSC phenotypic, transcriptional and secretory immunomodulatory profiles compared to 2D cultures. Further, CD146+ IFP-MSC spheroids showed enhanced secretory and transcriptional profiles, however, not reflected in a superior capacity to suppress activated PBMC suggesting 3D environment sufficient to induce an immunomodulatory phenotype. Crude IFP-MSC spheroids modulated the molecular response of synoviocytes previously exposed to inflammatory cues. Therapeutically, IFP-MSC spheroids retained Substance P degradation potential in vivo, while effectively induced resolution of inflammation/fibrosis of synovium and fat pad, halting the articular cartilage degradation in a rat model of progressive synovitis, fat pad fibrosis and osteoarthritis. Conclusions: 3D spheroids confer IFP-MSC a reproducible and enhanced immunomodulatory effect in vitro and in vivo, circumventing the requirement of non-compliant cell priming or selection before administration, thus streamlining cell products manufacturing protocols.