Efferocytosis of viable versus heat-inactivated MSC induces human monocytes to distinct immunosuppressive phenotypes.

BACKGROUND: Immunomodulation by mesenchymal stromal cells (MSCs) can occur through trophic factor mechanisms, however, intravenously infused MSCs are rapidly cleared from the body yet a potent immunotherapeutic response is still observed. Recent work suggests that monocytes contribute to the clearance of MSCs via efferocytosis, the body's natural mechanism for clearing dead and dying cells in a non-inflammatory manner. This begs the questions of how variations in MSC quality affect monocyte phenotype and if viable MSCs are even needed to elicit an immunosuppressive response. METHODS: Herein, we sought to dissect MSC's trophic mechanism from their efferocytic mechanisms and determine if the viability of MSCs prior to efferocytosis influences the resultant phenotype of monocytes. We cultured viable or heat-inactivated human umbilical cord MSCs with human peripheral blood mononuclear cells for 24 h and observed changes in monocyte surface marker expression and secretion profile. To isolate the effect of efferocytosis from MSC trophic factors, we used cell separation techniques to remove non-efferocytosed MSCs before challenging monocytes to suppress T-cells or respond to inflammatory stimuli. For all experiments, viable and heat-inactivated efferocytic-licensing of monocytes were compared to non-efferocytic-licensing control. RESULTS: We found that monocytes efferocytose viable and heat-inactivated MSCs equally, but only viable MSC-licensed monocytes suppress activated T-cells and suppression occurred even after depletion of residual MSCs. This provides direct evidence that monocytes that efferocytose viable MSCs are immunosuppressive. Further characterization of monocytes after efferocytosis showed that uptake of viable-but not heat inactivated-MSC resulted in monocytes secreting IL-10 and producing kynurenine. When monocytes were challenged with LPS, IL-2, and IFN-γ to simulate sepsis, monocytes that had efferocytosed viable MSC had higher levels of IDO while monocytes that efferocytosed heat inactivated-MSCs produced the lowest levels of TNF-α. CONCLUSION: Collectively, these studies show that the quality of MSCs efferocytosed by monocytes polarize monocytes toward distinctive immunosuppressive phenotypes and highlights the need to tailor MSC therapies for specific indications.

Toxicity Impacts on Human Adipose Mesenchymal Stem/Stromal Cells Acutely Exposed to Aroclor and Non-Aroclor Mixtures of Polychlorinated Biphenyl.

Polychlorinated biphenyl (PCB) accumulates in adipose where it may impact the growth and function of cells within the tissue. This is particularly concerning during adolescence when adipocytes expand rapidly. Herein, we sought to understand how exposure to PCB mixtures found in U.S. schools affects human adipose mesenchymal stem/stromal cell (MSC) health and function. We investigated how exposure to Aroclor 1016 and Aroclor 1254, as well as a newly characterized non-Aroclor mixture that resembles the PCB profile found in cabinets, Cabinet Mixture, affects adipose MSC growth, viability, and function in vitro. We found that exposure to all three mixtures resulted in two distinct types of toxicity. At PCB concentrations >20 µM, the majority of MSCs die, while at 1-10 µM, MSCs remained viable but display numerous alterations to their phenotype. At these sublethal concentrations, the MSC rate of expansion slowed and morphology changed. Further assessment revealed that PCB-exposed MSCs had impaired adipogenesis and a modest decrease in immunosuppressive capabilities. Thus, exposure to PCB mixtures found in schools negatively impacts the health and function of adipose MSCs. This work has implications for human health due to MSCs' role in supporting the growth and maintenance of adipose tissue.