pSTAT5-SOCS1 signalling as a novel pathway in macrophage metabolic reprogramming by Mesenchymal Stromal Cells (MSCs) in ARDS

Background: For effective clinical translation of MSC based therapies in ARDS better understanding of their mechanism of action is needed.  Macrophages are important cellular mediators of MSC immunomodulatory effect. Previously we showed that MSC metabolically reprogram macrophages towards anti-inflammatory phenotype through mitochondrial transfer in extracellular vesicles, but macrophage intracellular signalling pathways were not investigated. Here we evaluated the role of suppressor of Cytokine Signalling 1 (SOCS1) protein and Signal Transducer and Activator of Transcription 5(STAT5) axis in MSC effect on human macrophage reprogramming. Methods: Human monocyte-derived macrophages (MDMs) were stimulated with LPS or plasma samples from ARDS patients and treated with MSC conditioned medium (CM) or MSC-extracellular vesicles (EVs). SOCS1 and STAT5 protein expression was tested by Western blot, cytokine levels (TNF-a, IL-8) were measured by ELISA. Specific siRNAs or pharmacological inhibitors were used to silence SOCS1/STAT5 expression in MDMs. To evaluate the role of oxidative phosphorylation, MDMs were treated with Oligomycin. Results: MDM SOCS1 expression was downregulated by LPS and restored by MSC-CM. SOCS1 upregulation was dependent on STAT5 phosphorylation. SOCS1 silencing and pSTAT5 inhibition in MDMs impaired ability of MSC CM to suppress macrophage secretion of pro-inflammatory cytokines. These effects were recapitulated by MSC EVs and required macrophage oxidative phosphorylation, suggesting that mitochondrial transfer via EVs might be involved in the effect. EVs also upregulated SOCS1 and pSTAT5 expression when MDMs were incubated in plasma from ARDS patients. Conclusion: pSTAT5-SOCS1 signalling is a novel pathway involved in MSC macrophage reprogramming in ARDS.