Farida Djouad

Mesenchymal stem cells: innovative therapeutic tools for rheumatic diseases

Mesenchymal stem cells (MSCs), or multipotent mesenchymal stromal cells as they are also known, have been identified in bone marrow as well as in other tissues of the joint, including adipose, synovium, periosteum, perichondrium, and cartilage. These cells are characterized by their phenotype and their ability to differentiate into three lineages: chondrocytes, osteoblasts and adipocytes. Importantly, MSCs also potently modulate immune responses, exhibit healing capacities, improve angiogenesis and prevent fibrosis. These properties might be explained at least in part by the trophic effects of MSCs through the secretion of a number of cytokines and growth factors. However, the mechanisms involved in the differentiation potential of MSCs, and their immunomodulatory and paracrine properties, are currently being extensively studied. These unique properties of MSCs confer on them the potential to be used for therapeutic applications in rheumatic diseases, including rheumatoid arthritis, osteoarthritis, genetic bone and cartilage disorders as well as bone metastasis.

Transcriptional profiles discriminate bone marrow-derived and synovium-derived mesenchymal stem cells

Previous studies have reported that mesenchymal stem cells (MSC) may be isolated from the synovial membrane by the same protocol as that used for synovial fibroblast cultivation, suggesting that MSC correspond to a subset of the adherent cell population, as MSC from the stromal compartment of the bone marrow (BM). The aims of the present study were, first, to better characterize the MSC derived from the synovial membrane and, second, to compare systematically, in parallel, the MSC-containing cell populations isolated from BM and those derived from the synovium, using quantitative assays. Fluorescent-activated cell sorting analysis revealed that both populations were negative for CD14, CD34 and CD45 expression and that both displayed equal levels of CD44, CD73, CD90 and CD105, a phenotype currently known to be characteristic of BM-MSC. Comparable with BM-MSC, such MSC-like cells isolated from the synovial membrane were shown for the first time to suppress the T-cell response in a mixed lymphocyte reaction, and to express the enzyme indoleamine 2,3-dioxygenase activity to the same extent as BM-MSC, which is a possible mediator of this suppressive activity. Using quantitative RT-PCR these data show that MSC-like cells from the synovium and BM may be induced to chondrogenic differentiation and, to a lesser extent, to osteogenic differentiation, but the osteogenic capacities of the synovium-derived MSC were significantly reduced based on the expression of the markers tested (collagen type II and aggrecan or alkaline phosphatase and osteocalcin, respectively). Transcription profiles, determined with the Atlas Human Cytokine/Receptor Array, revealed discrimination between the MSC-like cells from the synovial membrane and the BM-MSC by 46 of 268 genes. In particular, activin A was shown to be one major upregulated factor, highly secreted by BM-MSC. Whether this reflects a different cellular phenotype, a different amount of MSC in the synovium-derived population compared with BM-MSC adherent cell populations or the impact of a different microenvironment remains to be determined. In conclusion, although the BM-derived and synovium-derived MSC shared similar phenotypic and functional properties, both their differentiation capacities and transcriptional profiles permit one to discriminate the cell populations according to their tissue origin.

The immunosuppressive signature of menstrual blood mesenchymal stem cells entails opposite effects on experimental arthritis and graft versus host diseases

Recently, a noninvasive and highly proliferative stem cell population from menstrual blood called MenSCs has been identified. Despite their use in clinical studies, their immunomodulatory properties have not yet been investigated. In this context, we studied the immunosuppressive properties of MenSCs in comparison with the well‐characterized bone marrow derived‐MSCs (BM‐MSCs). Using an in vitro proliferation assays, we showed that MenSCs displayed a lower suppressive effect on peripheral blood mononuclear cells and in particular on the proinflammatory CD4+IFN‐γ+ and CD8+IFNγ+ cells than BM‐MSCs. Moreover, compared to BM‐MSCs, MenSCs activated with IFN‐γ and IL‐1β produced lower amounts of immunosuppressive factors such as IDO, PDL‐1, PGE2, and Activin A and exhibited a substantial lower expression level of IFN‐γ receptor subunits. In the collagen induced arthritis model, while BM‐MSCs administration resulted in a potent therapeutic effect associated with a significant decrease of proinflammatory T cell frequency in the lymph nodes, MenSCs injection did not. In contrast, in the xeno‐GVHD model, only MenSCs administration significantly increased the survival of mice. This beneficial effect mediated by MenSCs was associated with a higher capacity to migrate into the intestine and liver and not to their anti‐inflammatory capacities. All together our results demonstrate for the first time that the therapeutic potential of MSC in the experimental xeno‐GVHD model is independent of their immunosuppressive properties. These findings should be taken into consideration for the development of safe and effective cell therapies. Stem Cells 2016;34:456–469

Involvement of Angiopoietin-like 4 in Matrix Remodeling during Chondrogenic Differentiation of Mesenchymal Stem Cells

Background: Due to their ability to differentiate into chondrocytes, mesenchymal stem cells (MSCs) are candidates for cartilage repair. Results: During chondrogenic differentiation of MSCs, angiopoietin-like 4 (ANGPTL4) triggers degradation and reduced synthesis of the cartilage matrix. Conclusion: ANGPTL4 promotes cartilage matrix remodeling. Significance: In the perspective of MSC-based cartilage engineering, inhibiting ANGPTL4 expression or action could help to stabilize cartilage formation. Mesenchymal stem cells (MSCs) are considered for cartilage engineering given their ability to differentiate into chondrocytes. Chondrogenic differentiation of MSCs is currently triggered by micromass culture in the presence of a member of the TGF-β superfamily. However, the main constituents of the cartilaginous matrix, aggrecan and type II collagen, are degraded at the end of the differentiation process through induction of matrix metallopeptidase (MMP)13. We hypothesized that MSCs undergoing chondrogenic differentiation produce an intermediate cytokine that triggers this matrix remodeling. Analysis of transcriptomic data identified angiopoietin-like 4 (ANGPTL4) as one of the most strongly up-regulated gene encoding a secreted factor during TGF-β-induced chondrogenesis. To gain insight into the role of ANGPTL4 during chondrogenesis, we used recombinant ANGPTL4 as well as a RNA interference approach. Addition of exogenous ANGPTL4 during the course of TGF-β-induced differentiation reduced the mRNA levels of aggrecan and type II collagen, although it increased those of MMP1 and MMP13. Accordingly, deposition of aggrecan and total collagens was diminished, whereas release of MMP1 and MMP13 was increased. Conversely, transfection of MSCs with an siRNA targeting ANGPTL4 prior to induction of chondrogenesis increased expression of type II collagen and aggrecan, whereas it repressed that of MMP1, MMP3, and MMP13. A neutralizing antibody against integrin αVβ5, a known receptor for ANGPTL4, mimicked some of the effects observed after siRNA-mediated ANGPTL4 silencing. Our data provide evidence that ANGPTL4 promotes cartilage matrix remodeling by inhibiting expression of its two key components and by up-regulating the level of certain MMPs.

Mesenchymal stem cells repress Th17 molecular program through the PD-1 pathway

MSC display potent suppressive properties initially described a decade ago. Since then, MSC suppressive activities on T-cell effector pathways have been highly investigated. MSC modulate CD4 cell differentiation through different mechanisms depending on culture conditions and display disparate activities on T cells according to their differentiation status. A significant amount of evidence for MSC effects on Th17 cells revealed that MSC could be suppressive under diverse circumstances but also enhance Th17 cell activity under other conditions. In the present study, we investigated the suppressive effects of MSC on Th1 and Th17 subsets of T cells using T cells undergoing Th1 and Th17 polarization or, on mature Th1 and Th17 cells. We showed that MSC inhibited the proliferation of mature Th1 cells and T cells during their differentiation toward Th1 cells. This suppressive effect was maintained in a transwell culture insert, where MSC were plated in the insert, demonstrating the major role played by soluble factors. Using the transwell barrier, we observed that MSC decrease the number of T cells undergoing Th17 differentiation whereas they did not affect IL-17 production by mature Th17, demonstrating the need for cell contact for suppressing Th17 cell function. We observed a high level of PD-L1 expression on MSC co-cultured with differentiating or polarized Th1 and Th17 cells. Importantly, using neutralizing antibodies specific for PD-L1 and PD-1, we showed that the mechanisms by which MSC mediate Th17 cell repolarization depend on PD-L1 expression on MSC. Taken together, our results demonstrated a cell-to-cell contact depend mechanism in the selective immunosuppression of MSC on mature Th17 cells through up-regulation of PD-L1.