Journal: PLOS ONE

Article Title: Recovery after human bone marrow mesenchymal stem cells (hBM-MSCs)-derived extracellular vesicles (EVs) treatment in post-MCAO rats requires repeated handling

doi: 10.1371/journal.pone.0312298

Figure Lengend Snippet: (a–c) Characterization and quantification of human-bone marrow mesenchymal stem cell (hBM-MSC)-derived extracellular vesicles (EVs). (a) Nanosight Tracking analyses (NTA) showing the size distribution pattern of EVs. (b) Western blots of hBM-MSCs marker (GM130) and EVs markers (ALIX, CD9, CD81). “Wash” lane represents the negative control. Each image comes from different blots. (c) Representative image of EVs using transmission electron microscopy (TEM). Scale bar 100 nm. (d–f) In vivo tracking of ExoGlow-labeled EVs demonstrated that EVs reach the brain and accumulate near the stroke injury 6 hours after administration at 2 dps. (d) Timeline of intranasal single dose treatment of ExoGlow-labeled EVs (~2.4 x 10 9 EVs in 200 μl), starting 2 dps and collecting the tissue 6 hours after. (e) Representative coronal section after a single intranasal administration of ExoGlow-labeled EVs (~2.4 x 10 9 EVs in 200 μl) at 2 dps. Hoechst indicates cell nuclei. Scale bars 4 mm and 100 μm. (f) Quantification of the average size of ExoGlow+ particles in the ipsilateral and contralateral hemispheres. Data was analyzed using a paired t-test (*p<0.05). Bars show mean +/− SD. Each symbol represents the average size of ExoGlow+ particles from all ROIs in each hemisphere (5 ROIs/hemisphere in each rat) of a single rat ( n = 4 ).

Article Snippet: Human bone marrow mesenchymal stem cells (hBM-MSCs) were purchased from Celprogren (frozen vial with ~1.2 x 10 6 cells; #36094–22) and plated on human mesenchymal bone marrow stem cell culture extracellular expansion matrix pre-coated T75 Flasks (Celprogen, #E36094-21-T75) following Celprogren recommendations.

Techniques: Derivative Assay, Western Blot, Marker, Negative Control, Transmission Assay, Electron Microscopy, In Vivo, Labeling

Journal: PLoS ONE

Article Title: Synaptically-Competent Neurons Derived from Canine Embryonic Stem Cells by Lineage Selection with EGF and Noggin

doi: 10.1371/journal.pone.0019768

Figure Lengend Snippet: Canine ESCs derived from whole embryo explants cultured on MEFs in the presence of LIF and bFGF exhibit tightly packed, flat human ESC-like colonies (A, B), and express OCT4 (C), SOX2 (D), ESC surface markers SSEA 3 (E), SSEA4 (F), Tra-1-81 (G) and FGFR2 (H). Expression of ESC makers was not seen in MEF feeder cells (arrowhead, C–H). Upon removal of LIF/bFGF and addition of 10% knock-out serum, cESC colonies rapidly differentiate (I) with outgrowths of radial cells at weeks 2–3 (J), which produce neural morphologies at 4 weeks (K) that express neural-specific protein MAP2 (L). DAPI was used as nuclear counterstain (blue). Scale bar = 100 µm. Abbreviations: cESC, canine embryonic stem cell; MEF, mouse embryonic fibroblast; Wk, week.

Article Snippet: Briefly, cESC lines produced from immunodissection of the ICM (OVC.ID) or explant of whole blastocysts (OVC.EX) were plated on 10 5 cells/cm 2 mitotically-arrested murine embryonic fibroblasts (MEF) (ATCC, SCRC-1040) in ESC media that consisted of knock-out (KO)-DMEM, 15% KO serum replacement, 4 ng/mL bFGF (Invitrogen), 10 ng/mL hrLIF (Sigma), 15 μM adenosine, cytidine, guanosine and uridine, 5 μM thymidine, 2 mM L-glutamine, 1× non-essential amino acids, and 0.1 mM beta-mercaptoethanol.

Techniques: Derivative Assay, Cell Culture, Expressing, Knock-Out

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Soluble RANKL cleaved from activated lymphocytes by TNF-α converting enzyme (TACE) contributes to osteoclastogenesis in periodontitis

doi: 10.4049/jimmunol.1601114

Figure Lengend Snippet: (A) RNAs from RAW 264.7 cells cultured with normal culture media (C), sRANKL (R; 100 ng/ml), TNF-α (T; 10 ng/ml), or 50% human GMC conditioned medium (sup.) were extracted after incubation for 5 days. The isolated RNAs were then reverse transcribed, and five osteoclast marker gene transcripts (ATP6v0d2, c-src, DC-STAMP, integrin β3, and vATPase) were detected by real-time RT-PCR. GAPDH was used as internal control. Fold increase from control in each transcript is shown. All experiments were performed in triplicate wells for each group. Results are expressed as mean ± SD. * p<0.001 vs. control. RAW 264.7 cells were co-cultured on OsteoAssay plate (Fig. 6B) or Osteologic plate (Fig. 6C) with 50% human GMC conditioned medium harvested from GMC culture with or without anti-TACE antibody. In some conditions, anti-RANKL antibody was added to block sRANKL activity (Fig. 6C). Three human GMC supernatants were analyzed. C: cultured with normal culture media; R: cultured with sRANKL (100 ng/ml). All experiments were performed in triplicate wells for each group. Results are expressed as mean ± SD. *: p<0.05 vs. control. †: p<0.05 between groups.

Article Snippet: Pit formation assays To evaluate resorption activity of TRAP-positive multinucleated cells differentiated from RAW 264.7 cells, a resorption assay was performed using OsteoAssay TM Human Bone Plate (Corning, Corning, NY) or osteologic discs (BD Biosciences).

Techniques: Cell Culture, Incubation, Isolation, Reverse Transcription, Marker, Quantitative RT-PCR, Control, Blocking Assay, Activity Assay