Abstract
BackgroundRecently, it has been observed that mesenchymal stem cells (MSCs) can modulate their immunoregulatory properties depending on the specific in-vitro activation of different Toll-like receptors (TLR), such as TLR3 and TLR4. In the present study, we evaluated the effect of polyinosinic:polycytidylic acid (poly(I:C)) and lipopolysaccharide (LPS) pretreatment on the immunological capacity of MSCs in vitro and in vivo.MethodsC57BL/6 bone marrow-derived MSCs were pretreated with poly(I:C) and LPS for 1 hour and their immunomodulatory capacity was evaluated. T-cell proliferation and their effect on Th1, Th17, and Treg differentiation/activation were measured. Next, we evaluated the therapeutic effect of MSCs in an experimental autoimmune encephalomyelitis (EAE) model, which was induced for 27 days with MOG35–55 peptide following the standard protocol. Mice were subjected to a single intraperitoneal injection (2 × 106 MSCs/100 μl) on day 4. Clinical score and body weight were monitored daily by blinded analysis. At day 27, mice were euthanized and draining lymph nodes were extracted for Th1, Th17, and Treg detection by flow cytometry.ResultsPretreatment of MSCs with poly(I:C) significantly reduced the proliferation of CD3+ T cells as well as nitric oxide secretion, an important immunosuppressive factor. Furthermore, MSCs treated with poly(I:C) reduced the differentiation/activation of proinflammatory lymphocytes, Th1 and Th17. In contrast, MSCs pretreated with LPS increased CD3+ T-cell proliferation, and induced Th1 and Th17 cells, as well as the levels of proinflammatory cytokine IL-6. Finally, we observed that intraperitoneal administration of MSCs pretreated with poly(I:C) significantly reduced the severity of EAE as well as the percentages of Th1 and Th17 proinflammatory subsets, while the pretreatment of MSCs with LPS completely reversed the therapeutic immunosuppressive effect of MSCs.ConclusionsTaken together, these data show that pretreatment of MSCs with poly(I:C) improved their immunosuppressive abilities. This may provide an opportunity to better define strategies for cell-based therapies to autoimmune diseases.
Highlights
It has been observed that mesenchymal stem cells (MSCs) can modulate their immunoregulatory properties depending on the specific in-vitro activation of different Toll-like receptors (TLR), such as toll like receptor 3 (TLR3) and toll like receptor 4 (TLR4)
TLR3 and TLR4 pretreatment differentially affect the in-vitro immunosuppressive capacity of murine MSCs To evaluate the effect of specific stimulation of TLR3 and TLR4, we treated the MSCs for 1 hour with poly(I:C) or LPS in complete alpha minimal essential medium (αMEM) and determined their in-vitro immunomodulatory capacity
Flow cytometry was used to analyze the proliferation of CD3+ T lymphocytes that were cultured in the presence or absence of untreated MSCs or MSCs pretreated with poly(I:C) or LPS at different MSC:splenocyte ratios (1:5, 1:10, and 1:20), as described in Methods
Summary
It has been observed that mesenchymal stem cells (MSCs) can modulate their immunoregulatory properties depending on the specific in-vitro activation of different Toll-like receptors (TLR), such as TLR3 and TLR4. Mesenchymal stem cells (MSCs) are nonhematopoietic, multipotent progenitor cells isolated from a variety of adult tissues, including bone marrow and adipose tissue They are capable of self-renewal and are able to differentiate into at least some mesenchymal cell types, such as bone, cartilage, and fat, playing a potential role in tissue repair [1, 2]. In addition to their potential for differentiation, MSCs exhibit immunosuppressive activity, as shown by their ability to inhibit the proliferation and function of immunocompetent cells, such as T and B lymphocytes, natural killer cells, and dendritic cells [3,4,5]. We recently demonstrated that the intravenous administration of MSCs in EAE at different stages of the disease induced differential therapeutic effects depending on the proinflammatory environment at each stage of the disease [12]
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