Supplementary Materials Movie S1 Confocal video showing the Z stacks of demyelinated cerebellar slices treated with yMSC\CM for 8 days

Supplementary Materials Movie S1 Confocal video showing the Z stacks of demyelinated cerebellar slices treated with yMSC\CM for 8 days. in triplicate and one\way ANOVA\Tukey post hoc was used for statistical analysis. *Differences between passage 2 and passage 6; #differences between passage 2 and passage 4. **o ## the percentage of Ki67+ (a) and Caspase 3+ (b) cells were determined to evaluate cell proliferation and survival, respectively. No differences were observed in OPCs exposed to yMSC\CM compared cells incubated under control conditions. NS, not significant. GLIA-67-1510-s003.tif (6.4M) GUID:?4101C109-55CE-4980-9F28-57EE73A52FC1 Figure S3 Transplanted MSCs do not affect OPC activation and proliferation during CNS remyelination. Twelve months old rats were demyelinated by ethidium bromide injection into the caudal cerebellar peduncle. One, two, and three days after demyelination, yMSCs or oMSCs were systemically transplanted. PBS is used as vehicle control. Quantitative analysis shows the number of proliferating cells (Ki67+) (a) GYKI-52466 dihydrochloride and the number of activated proliferating OPCs (Nkx2.2+/Ki67) (b) within the lesion area (mm2) at 21 dpl. Note that none of transplanted groups shows significant difference in the number of proliferating cells and proliferating OPCs respect to the vehicle group. NS, not significant. GLIA-67-1510-s004.tif (6.4M) GUID:?D66526B1-B204-4F3F-8AF1-505B8521698D Figure S4 Transplanted MSCs do not transdifferentiate into cells from the oligodendroglial lineage during CNS remyelination. Twelve months old rats were demyelinated by ethidium bromide injection into the caudal cerebellar peduncle. One, two, and three days after demyelination, rats were systemically transplanted with yMSCs that express mCherry for their detection. Images show the presence of mCherry\expressing MSCs (red) as well as Olig2+ cells (green) at 21 dpl. Hoechst shows nuclei counterstaining (blue). GYKI-52466 dihydrochloride Dashed lines denote demyelinating lesion area (L). Scale bar?=?50?m. Inset in merge image shows a high magnification of the area limited by the square. Note that none of the transplanted mCherry\expressing MSCs coexpress the oligodendrocyte lineage marker Olig2. GLIA-67-1510-s005.tif (5.7M) GUID:?BFCF6DD1-E6FA-4CC7-9E99-A40FE805CD57 Figure S5 Periaxin\positive Schwann Cells do not coexpress the inflammatory cell marker Iba1 during CNS remyelination. Twelve months old rats were demyelinated by ethidium bromide injection into the caudal cerebellar peduncle. Left panel shows the presence of Periaxin\expressing Schwann cells (green) and Iba1+ positive inflammatory cells (red) at 21 dpl. Hoechst shows nuclei counterstaining (blue). Dashed lines denote demyelinating lesion area (L). Scale bar?=?100?m. Rest of the panels show a higher magnification of the area limited by the square. In merge image, the inset shows a magnification of the area delimited from the square. Note the absence of Periaxin\positive Schwann cells coexpressing the inflammatory cell marker Iba1. GLIA-67-1510-s006.tif (7.4M) GUID:?2C675992-F8A9-455E-8DC4-695763673AA6 Data Availability Statement Data Availability Statement: The data that support the findings of this study are available from your related author upon reasonable request. The data that support the findings of this study are available from your corresponding author upon reasonable request. Abstract Multiple sclerosis (MS) is definitely a demyelinating disease of the central nervous system (CNS) that leads to severe neurological deficits. Because of the immunomodulatory and neuroprotective activities and their ability to promote the generation of oligodendrocytes, mesenchymal stem cells (MSCs) are currently being developed for autologous cell therapy in MS. As ageing reduces the regenerative capacity of all cells, it is of relevance to investigate whether MSCs retain their pro\oligodendrogenic activity with increasing age. We demonstrate that MSCs Mouse monoclonal to CD38.TB2 reacts with CD38 antigen, a 45 kDa integral membrane glycoprotein expressed on all pre-B cells, plasma cells, thymocytes, activated T cells, NK cells, monocyte/macrophages and dentritic cells. CD38 antigen is expressed 90% of CD34+ cells, but not on pluripotent stem cells. Coexpression of CD38 + and CD34+ indicates lineage commitment of those cells. CD38 antigen acts as an ectoenzyme capable of catalysing multipe reactions and play role on regulator of cell activation and proleferation depending on cellular enviroment derived from aged rats have a reduced capacity to induce oligodendrocyte differentiation of adult CNS stem/progenitor cells. Ageing also abolished the ability of MSCs to enhance the generation of myelin\like sheaths in demyelinated cerebellar slice cultures. Finally, inside a rat model for CNS demyelination, ageing suppressed the capability of systemically transplanted MSCs to boost oligodendrocyte progenitor cell (OPC) differentiation during remyelination. Therefore, ageing restricts the ability of MSCs to support the generation of oligodendrocytes and consequently inhibits their capacity to enhance the generation of myelin\like GYKI-52466 dihydrochloride sheaths. These findings may impact on the design of therapies using autologous MSCs in older MS individuals. for 10 min and the cell pellet was resuspended in 1 mL MEM\10% FBS. About 250?L of each cell suspension was dissolved in CASYTON (Roche, Vienna Austria) and cell number was determined with CASY\Cell Counter (Roche, Vienna Austria). Measurements with CASY\Cell Counter were performed using the following settings: Dilution: 2.000e +00, Capillary: 150?m, and filtered through a 0.22?m\pore filter. 2.8. Bioluminescence assays Bioluminescence assays were performed using GYKI-52466 dihydrochloride noncommercial dual luciferase enzyme assay system. In this system,.