In vitro models for biological activity evaluation of a new antifibrotic drug based on the vesicular fraction of the human mesenchymal stromal cells secretome

Introduction. The vesicular fraction of human mesenchymal stem/stromal cells (EV MSCs) secretome can be considered as a pharmaceutical substance for the biological drug development to treat fibrotic diseases. Biological drugs quality control requires standard tests using simple validable methods. Additionally, it is necessary to assess specific activity using relevant models based on specific cellular targets and mechanisms of action.

Aim. Development of cell models for specific activity assessment of the biological drug from human EV MSCs for the fibrotic diseases treatment.

Materials and methods. We proposed two in vitro models to evaluate specific activity: the first one is primary human dermal fibroblast (HDF) cell lines induced differentiation model and the second one is polarization of macrophages derived from human peripheral blood monocytes.

Results and discussion. Transforming growth factor (TGFβ-1) treatment of HDF increased the level of the main myofibroblasts marker – alpha-smooth muscle actin (αSMA) within 96 hours. The simultaneous action of TGFβ-1 and EV-MSCs significantly decreased αSMA level compared to TGFβ-1-stimulated fibroblasts. Macrophages polarization towards M1-type with LPS/IFNγ combination resulted in increased IL-6, IL-12p35, TNFα genes expression after both 4 and 24 hours. The EV-MSCs addition to M1-type decreased the gene expression of proinflammatory cytokines IL-6, IL-12p35, TNFα in 24 hours.

Conclusion. We developed two in vitro models to assess specific activity of antifibrotic drug based on human EV-MSCs. In the first model value of specific activity is at least 2.5-fold decrease of αSMA level in TGFβ-1-stimulated HDF, comparing to non-treated control. In the second model the value is at least two-fold decrease in the level of IL-12p35, IL-6, TNFα expression in M1 macrophages, compared to non-treated M1 macrophages.

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