1. Allogeneic Mesenchymal Stem Cells for Treatment of Experimental Autoimmune Encephalomyelitis 
Moutih Rafei, Elena Birman, Kathy Forner, Jacques Galipeau 
Molecular Therapy 
Volume 17, Issue 10, Pages (October 2009)
DOI: /mt
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

2. Figure 1
Mesenchymal stromal cell (MSC) isolation. (a) MSC phenotypic analysis. Cultured C57Bl/6 MSCs were stained for various cell surface markers and shown to express CD44, CD73, CD105, and no apparent CD45. (b) MSC plasticity. As shown in b, MSC culture under adipogenic or osteogenic conditions leads to their differentiation.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

3. Figure 2
CCL2 characterization in MSC CM. (a) MSCs secrete CCL2. To prove that MSCs are capable of secreting the CCL2 chemokine, CM collected from NIH-3T3, or MSCs with or without IFN-γ (2 ng/ml) treatment were collected and tested by enzyme-linked immunosorbent assay. CCL2 was only detected in MSC CM and was upregulated following IFN-γ stimulation (n = 5 and *P < ). (b) MHCI and II were assessed by flow cytometry as indicative of successful IFN-γ treatment. CM, conditioned media; IFN-γ, interferon-γ; MHC, major histocompatibility complex; MSC, mesenchymal stromal cell.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

4. Figure 3
MSC-derived CCL2 and effects on mixed lymphocyte reaction (MLR). (a) MSC CM blocks two-way MLR in a dose-dependent manner. CM was collected from resting syngeneic or allogeneic MSCs and concentrated up to a 100×. A twofold serial dilution was then prepared and added to the MLR reaction. IFN-γ inhibition could only be achieved using a concentration ≥25× (n = 5 and *P < ). (b) MSC-derived CCL2 blocks IFN-γ during MLR. To demonstrate the direct involvement of CCL2 in IFN-γ inhibition, the addition of CCL2 neutralizing antibody (2.5 µg/ml) to the MLR reaction has reversed the IFN-γ inhibition effect to a comparable level with the positive control (n = 5 and *P < ). CM, conditioned media; IFN-γ, interferon-γ; MSC, mesenchymal stromal cell; n.d., not detected; NS, not significant.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

5. Figure 4
Therapeutic effects of syngeneic and allogeneic MSCs in experimental autoimmune encephalomyelitis (EAE). (a) Syngeneic and allogeneic MSC effects on EAE. Mice (n = 10/condition) with developed EAE symptoms were injected intraperitoneally at two different time points as shown by arrows (2 × 106 cells). Both syngeneic and allogeneic MSCs can lead to EAE improvement. (b) IFN-γ pretreatment of allogeneic MSCs leads to no therapeutic effects. The same in vivo EAE experiment as in (a) was repeated but comparing allogeneic MSCs with or without IFN-γ (2 ng/ml) pretreatment in vitro. No therapeutic improvements were noticed in the IFN-γ pretreated allogeneic MSCs compared to the PBS control group. IFN-γ, interferon-γ; MSC, mesenchymal stromal cell; PBS, phosphate-buffered saline.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

6. Figure 5
Allogeneic MSCs decreases the levels of pro-inflammatory cytokines. (a) IFN-γ and interleukin-17 (IL-17) levels in systemic circulation. Only allogeneic MSCs were capable of decreasing the circulating levels of IFN-γ and IL-17 in experimental autoimmune encephalomyelitis (EAE) mice, whereas IFN-γ pretreatment of allogeneic MSCs leads to no difference as compared to the PBS control group (n = 5 and *P < ). (b) MSCs tolerogenic effects. To demonstrate the tolerogenic effects of MSCs toward allogeneic cells, splenocytes collected from EAE mice injected with allogeneic MSCs were stimulated in vitro with the same MSCs they have been injected with. A robust decrease in IFN-γ secretion was obtained with mice treated with allogeneic MSCs, whereas MSC pretreatment with IFN-γ prior to their injection leads to no difference in comparison to the PBS control group. IFN-γ, interferon-γ; MSC, mesenchymal stromal cell; PBS, phosphate-buffered saline.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions

7. Figure 6
CD4 T-cell infiltration of the spinal cord. (a) Spinal cords of experimental autoimmune encephalomyelitis (EAE) mice were stained with H&E to demonstrate the levels of immune cells. As demonstrated in a, no differences could be seen between the PBS EAE mice or EAE mice injected with IFN-γ pretreated MSC. (b) The same outcome was obtained when CD4 T-cell infiltrates were assessed by flow cytometry (P < 0.05). IFN-γ, interferon-γ; H&E, hematoxylin and eosin; MSC, mesenchymal stromal cell; PBS, phosphate-buffered saline.
Molecular Therapy  , DOI: ( /mt )
Copyright © 2009 The American Society of Gene & Cell Therapy Terms and Conditions