Additional file 2: of Targeted intracerebral delivery of the anti-inflammatory cytokine IL13 promotes alternative activation of both microglia and macrophages after stroke

Figure S2. Representative example of MSC and IL13-MSC graft site remodeling within the MCAO brain lesion site. Control MSC (upper panel) and IL13-MSC (lower panel) grafts are able to survive in the pro-inflammatory stroke environment and display a similar remodeling pattern. MSC graft-infiltrating CCR2RFP/+ monocytes/macrophages (in red) at the core of the MSC grafts and brain-resident CX3CR1eGFP/+ microglia (in green) and astrocytes (in blue, first row) surrounding the MSC grafts. Arginase1 expression (in blue, second and third row) and Ym1 expression (in magenta, third row) by microglia and monocytes/macrophages, as a direct result of stimulation by IL13, was only detected in IL13-MSC grafts, but not in control MSC grafts. Scale bar 100 Οm. (TIF 13199 kb

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-4

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>ed slides pre-and post-LCM. (B) Upper graph showing the level of transgenic EGFP mRNA transcripts/1000 copies GAPDH in hMSC-EGFP cell cultures (CELLS, n = 1), on transplanted hMSC-EGFP cells at day 1 post-transplantation (DAY 1, n = 1), and on transplanted hMSC-EGFP cells at day 7 post-transplantation (DAY 7, n = 1). Lower graph showing the level of transgenic EGFP mRNA transcripts/1000 copies GAPDH in hMSC-NT3-EGFP cell cultures (CELLS, n = 1), on transplanted hMSC-NT3-EGFP cells at day 1 post-transplantation (DAY 1, n = 1), and on transplanted hMSC-NT3-EGFP cells at day 7 post-transplantation (DAY 7, n = 1)

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-5

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>hematoxylin-eosin (HE) staining indicating the presence of nucleated cells into the subcutaneously transplanted matrigel. Second row: direct EGFP fluorescence indicating the presence of EGFP positive hMSC-NT3-EGFP cell transplants. Third row: immuno-histochemical staining for EGFP indicating the presence of EGFP positive hMSC-NT3-EGFP cell transplants. Fourth row: immuno-histochemical staining for human mitochondrial antigen (hMA) indicating the human nature of the observed EGFP positive hMSC-NT3-EGFP cell transplants. Representative pictures were chosen from multiple subcutaneous hMSC-NT3-EGFP cell transplants analysed 1 day (n = 6), 1 week (n = 5) and 2 weeks (n = 3) post-transplantation

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-7

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>mals (+ cyclosporine A). Right side: histological assessment of tumour regression in rat spinal cord at week 4 post-transplantation upon withdrawal of immune suppression at week 2 post-transplantation (-cyclosporine A). First row: hematoxylin-eosin (HE) staining indicating localisation and general appearance of transplantation site. Second row: direct EGFP fluorescence indicating the presence or absence of EGFP positive hMSC-EGFP cell transplants. Third row: immuno-histochemical staining for EGFP indicating the presence or absence of EGFP positive hMSC-EGFP cell transplants. Fourth row: immuno-histochemical staining for CD68 indicating macrophage infiltration into the transplantation site. Representative pictures were chosen from multiple hMSC-EGFP cell transplanted spinal cords analysed 1 day (n = 4), 2 weeks (n = 3) and 4 weeks (n = 2) post-transplantation

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-0

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>pulations. CMV: Cytomegalovirus immediate early promotor + enhancer. EGFP: enhanced green fluorescent protein. pA: SV40 early mRNA polyadenylation signal. NT3: neurothrophin-3. IRES: internal ribosome entry site. (B) Representative standard PCR and RT-PCR analysis on DNA and mRNA isolated from the different genetically modified hMSC populations used in this study (see numbers below pictures) indicating the presence of transgenic EGFP and/or NT3 DNA and mRNA sequences. M: length marker. GAPDH: glyceraldehyde-3-phosphate dehydrogenase. (C) Representative real-time RT-PCR analysis on mRNA isolated from the different genetically modified hMSC populations used in this study (see numbers below pictures) showing quantitative differences in the level of transgenic EGFP and/or NT3 mRNA transcripts/1000 copies GAPDH; nd: no data available. (D) Representative ELISA measurement on supernatant samples from the different genetically modified hMSC populations used in this study (see numbers below pictures) showing quantitative differences in the level of NT3 secretion in picogram/10cells/24 hours. (E) Representative flow cytometric analysis of EGFP expression by hMSC-EGFP and hMSC-NT3-EGFP populations showing quantitative differences in the level of transgenic EGFP protein expression. SSC: side scatter

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-1

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>d week 1–4). First row: hematoxylin-eosin (HE) staining indicating localisation and general appearance of transplantation site. Second row: direct EGFP fluorescence indicating the presence or absence of EGFP positive hMSC-EGFP cell transplants. Third row: immuno-histochemical staining for EGFP indicating the presence or absence of EGFP positive hMSC-EGFP cell transplants. Fourth row: immuno-histochemical staining for CD68 indicating macrophage infiltration into the transplantation site. All slides were examined using a conventional light/fluorescence microscope and digital pictures were taken under magnification as indicated. Representative pictures were chosen from multiple hMSC-EGFP cell transplanted spinal cords analysed for day 1 (n = 2), week 1 (n = 2), week 2 (n = 2), week 3 (n = 2), and week 4 (n = 6) post-transplantation. (B) Molecular and histological assessment of hMSC-EGFP cell transplant survival in rat spinal cord under systemic immune suppression (subcutaneous 10 mg/kg/day cyclosporin A) at four time points post transplantation (day 1 and week 1–3). DAY 1: Real-time RT-PCR analysis on mRNA isolated from hMSC-EGFP cell transplanted spinal cords on day 1 post-transplantation (n = 2) indicating the presence of EGFP mRNA transcripts in spinal cord following hMSC-EGFP cell transplantation. WEEK 1: direct EGFP fluorescence and immuno-histochemical staining for EGFP indicating the presence of EGFP positive hMSC-EGFP cell transplants on week 1 post-transplantation. Representative pictures were chosen from multiple hMSC-EGFP cell transplanted spinal cords analysed 1 week post-transplantation (n = 5). WEEK 2: HE staining, direct EGFP fluorescence and immuno-histochemical staining for EGFP indicating the presence of EGFP positive hMSC-EGFP cell transplants on week 2 post-transplantation. Representative pictures were chosen from multiple hMSC-EGFP cell transplanted spinal cords analysed 2 weeks post-transplantation (n = 3). WEEK 3: Standard PCR and RT-PCR analysis on DNA and mRNA isolated from hMSC-EGFP cell-transplanted spinal cords 3 weeks post-transplantation indicating the presence of EGFP DNA sequences and EGFP mRNA transcripts in spinal cord following hMSC-EGFP cell transplantation

Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord-2

<p><b>Copyright information:</b></p><p>Taken from "Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord"</p><p>http://www.biomedcentral.com/1472-6750/7/90</p><p>BMC Biotechnology 2007;7():90-90.</p><p>Published online 14 Dec 2007</p><p>PMCID:PMC2225398.</p><p></p>spinal cords on day 1 post-transplantation (n = 2) and on week 1 post-transplantation (n = 2) demonstrating the absence of detectable exogenous NT3 mRNA transcripts. SC: spinal cord