Time-dependent LPS exposure commands MSC immunoplasticity through TLR4 activation leading to opposite therapeutic outcome in EAE

Abstract

Background Mesenchymal stem cells (MSCs) have been recognized for their regenerative and anti-inflammatory capacity which makes them very attractive to cell therapy, especially those ones to treat inflammatory and autoimmune disease. Two different immune-phenotypes have been described for MSCs depending on which Toll-like receptor (TLR) is activated. MSC1 is endowed with a pro-inflammatory phenotype following TLR4 activation with LPS. On the other hand, anti-inflammatory MSC2 is induced by the activation of TLR3 with Poly(I:C). High immunoplasticity of MSCs is a matter of concern in cell-based therapies. In this study, we investigated whether a single stimulus can induce both types of MSCs through a differential activation of TLR4 with LPS. Methods MSCs were activated with LPS following a short exposure of 1-h (MSCs-LPS1h) or long-time exposure for 48 h (MSCs-LPS48h), and then, we evaluated the biological response in vitro, the immunosuppressive capacity of MSCs in vitro, and the therapeutic potential of MSCs in an experimental autoimmune encephalomyelitis (EAE) mouse model. Results Our results showed that 1-h LPS exposure induced a MSC1 phenotype. Indeed, MSCs-LPS1h expressed low levels of NO/iNOS and decreased immunosuppressive capacity in vitro without therapeutic effect in the EAE model. In contrast, MSCs-LPS48h achieved a MSC2-like phenotype with significant increase in the immunosuppressive capacity on T cell proliferation in vitro, together with an improved in the therapeutic effect and higher Treg, compared to unstimulated MSCs. Furthermore, we determine through the MSCs-TLR4KO that the expression of TLR4 receptor is essential for MSCs’ suppressive activity since TLR4 deletion was associated with a diminished suppressive effect in vitro and a loss of therapeutic effect in vivo. Conclusions We demonstrate that MSCs display a high immunoplasticity commanded by a single stimulus, where LPS exposure time regulated the MSC suppressive effect leading into either an enhanced or an impairment therapeutic activity. Our results underscore the importance of phenotype conversion probably related to the TLR4 expression and activation, in the design of future clinical protocols to treat patients with inflammatory and autoimmune diseases.

Effects of bacterial lipopolysaccharide and Shiga Toxin on induced Pluripotent Stem Cell-derived Mesenchymal Stem Cells

“…In this sense, in the proteomic analysis we found that the use of LPS+Stx in iPSC-MSC, in-duced the release of proteins related to the BP of "IL-12 mediated signalling pathway" such as PPIA, which is reported to promote apoptosis in endothelial cells and chemotaxis in inflammatory cells [36]. As Wong and Waterman et.al described in a previous work, we can speculate that activation of TLR4 in iPSC-MSC due to LPS resulted in the secretion of pro-inflammatory factors in the CM that are important for early injury responses, like migration and cell adhesion to ECM, but not to resolve tissue damage [9]. However, these mechanisms could prepare the microenvironment for later iPSC-MSC anti-inflammatory responses that could facilitate the restoration of tissue injury.…”

Mesenchymal Stem Cells Can Both Enhance and Inhibit the Cellular Response to DNA Immunization by Genes of Nonstructural Proteins of the Hepatitis C Virus

“…MSC1 is endowed with a proinflammatory phenotype following TLR4 activation with LPS. On the other hand, anti-inflammatory MSC2 is induced by the activation of TLR3 with Poly(I:C) [ 55 , 56 , 57 ]. MSC1 shows an increased synthesis and secretion of proinflammatory cytokines and chemokines, such as IL-6 and IL-8, whereas MSC2 has increased production of immunosuppressive mediators, such as IP-10 and CCL5.…”

“…MSC1 shows an increased synthesis and secretion of proinflammatory cytokines and chemokines, such as IL-6 and IL-8, whereas MSC2 has increased production of immunosuppressive mediators, such as IP-10 and CCL5. MSCs have high immunoplasticity, and the phenotype conversion can be caused by exogenous stimuli, such as proinflammatory cytokines or TLR agonists, as well as the duration of treatment [ 15 , 57 , 58 ]. The data on the effect of exogenous proinflammatory cytokines on MSCs are contradictory.…”

A Chemically Defined, Xeno- and Blood-Free Culture Medium Sustains Increased Production of Small Extracellular Vesicles From Mesenchymal Stem Cells

“…A few dozen cell-based therapies have obtained market authorization in several countries ( Cuende et al, 2018 ), and at least another dozen of approved MSC-based therapies are expected to reach the market by the year 2030 ( Olsen et al, 2018 ). We have recently demonstrated ( Kurte et al, 2020 ) that MSC’s high immunoplasticity depends on the exposure duration with the inflammatory milieu, leading into either an enhanced or an impairment therapeutic activity, a matter of great concern for their clinical use. Both the translational advances and the limitation of the use of MSC in some clinical applications have pushed the field toward exploring their therapeutic potential without the need for cell transplantation.…”