Abstract
Introduction: To investigate the influence of secretomes from native and dexamethasone-treated adipose-derived multipotent mesenchymal stromal cells (MMSC) on the proliferation of mononuclear cells (MNCs) and on their expression of phospho-NF-κB p65 in vitro.
 Materials and Methods: MMSCs were isolated from the fat of 5 healthy donors. The cells were grown in culture up to passage four, then treated with dexamethasone for 3 hours, washed off the preparations, and incubated in a serum-free medium for 48 hours. Some of the cells were not treated with dexamethasone. Supernatants from cell cultures were concentrated by ultrafiltration, standardized by the content of galectin-1, sterilized, and added to MNCs from peripheral blood of 8 healthy donors. MNCs were isolated in a Ficoll density gradient according to a standard protocol. The expression of phospho-NF-κB p65 and Ki-67 in MNCs under the influence of MMSC secretomes in isotypic and negative controls was determined on a flow cytometer.
 Results and Discussion: The expression of phospho-NF-kκB p65 and Ki-67 is decreased by the MMSC secretome. At the same time, a statistically significant decrease in phospho-NF-κB p65 by 36.2% (p< 0.05) is observed when using a secretome from native cells. Ki-67 expression is reduced by 42.3% (p < 0.05) under the influence of a secretome from dexamethasone-treated MMSCs.
 Conclusion: The MMSC secretome, as well as MMSCs themselves, has an anti-inflammatory effect due to the effect on the expression of the active form of NF-κB and the proliferative activity of mononuclear cells. At the same time, pretreatment of cells with dexamethasone reduces the effect on phospho-NF-κB expression and increases the inhibitory effect on MNC proliferation.
Highlights
To investigate the influence of secretomes from native and dexamethasone-treated adipose-derived multipotent mesenchymal stromal cells (MMSC) on the proliferation of mononuclear cells (MNCs) and on their expression of phospho-NF-κB p65 in vitro
Inhibition of NF-κB using dexamethasone regulates the intracellular domain of the Notch signaling pathway, which reduces macrophage differentiation, and it is a potential target for intervention, for example, in atherosclerosis (Binesh et al 2019)
The authors showed a decrease in NF-κB expression and an increase in IL-10 expression when dexamethasone was previously injected to mice before stimulating lung inflammation with LPS (Al-Harbi et al 2016)
Summary
To investigate the influence of secretomes from native and dexamethasone-treated adipose-derived multipotent mesenchymal stromal cells (MMSC) on the proliferation of mononuclear cells (MNCs) and on their expression of phospho-NF-κB p65 in vitro. The expression of phospho-NF-κB p65 and Ki-67 in MNCs under the influence of MMSC secretomes in isotypic and negative controls was determined on a flow cytometer. Pretreatment of cells with dexamethasone reduces the effect on phospho-NF-κB expression and increases the inhibitory effect on MNC proliferation. Intracellular marker Ki-67 was chosen to study the effect of the MMSC secretome on the proliferative activity of MNCs, because Ki-67 is expressed only in proliferating cells and plays a role in early stages of rRNA synthesis. Intracellular marker Ki-67 was chosen to study the effect of the MMSC secretome on the proliferative activity of MNCs, because Ki-67 is expressed only in proliferating cells and plays a role in early stages of rRNA synthesis. (Bullwinkel et al 2006)
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