Journal: Life science alliance

Article Title: Megakaryocytes possess a STING pathway that is transferred to platelets to potentiate activation.

doi: 10.26508/lsa.202302211

Figure Lengend Snippet: Figure 1. STING stimulation of megakaryocytes induces a type-I interferon response. (A) Brightfield images of megakaryocytes isolated using anti-CD41 magnetic beads at day 4 of in vitro differentiation. Enlargement shows the production of proplatelets (black arrows) by mature megakaryocytes. Scale bar: 30 µm. (B, C) Cytokine production analysis by ELISA of megakaryocytes stimulated with DMXAA for 5 h (B) and 24 h (C) showing a significant increase in IFNB and CCL5 cytokines, n = 4 independent experiments; each dot represents data from one independent experiment. The graph shows data ± STD, and data were analysed by unpaired two-tailed t tests, *P < 0.05. (D, E) Representative immunofluorescence images of megakaryocytes taken by confocal microscopy showing the localisation of cGAS or STING (red), ERp57 or lamin A/C (green), and chromatin (blue), representative of n = 3 independent experiments. Scale bars: 20 µm (D) and 15 µm (E). (F) Representative immunofluorescence images of CD41-positive (magenta for CD41+ and blue for DNA) and CD41-negative (only blue) cells taken by confocal microscopy. Representative images of more than n = 3 independent experiments. Scale bars: 30 µm.

Article Snippet: Cells were then labelled with 10 μl anti-CD41 antibody (coupled to Biotin or APC—Miltenyi Biotec) for 10 min at 4°C, then washed with 2 ml of isolation buffer, and centrifuged at 300g for 10 min at 4°C, twice before the addition of 20 μl of Anti-Biotin/APC MicroBeads (Miltenyi Biotec) and 70 μl of isolation buffer for 15 min at 4°C.

Techniques: Isolation, Magnetic Beads, In Vitro, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Confocal Microscopy

Journal: International journal of molecular sciences

Article Title: Pulmonary Biodistribution of Platelet-Derived Regenerative Exosomes in a Porcine Model.

doi: 10.3390/ijms25052642

Figure Lengend Snippet: Figure 1. Exosome characterization of PEP and membrane labeling by lipophilic dye. (A) NanoSight analysis of PEP demonstrating distribution of particle size ± standard deviation (gray shading). (n = 3). (B) Transmission electron microscopy image of PEP with scale bar 2 µm. (C) Western blot demonstrating presence of hallmark exosomal proteins CD9, CD63, and Flotillin, as well as platelet integrin CD41. (D) Immunocytochemistry of human umbilical vein endothelial cells following treatment with DiI-labeled PEP or DiI alone (control) in red, counterstained with phalloidin (green) and DAPI (blue), scale bar 20 µm. (E) Quantification (n = 5) of mean integrated density DiI per nuclei (±SD) for Figure 1D. ** p < 0.001 using Mann-Whitney two-tailed t-test. (F) Xenogen image demonstrating fluorescent signal detected in unlabeled PEP, DiI, and PEP + DiI. (G) Xenogen image demonstrating fluorescent signal detected in unlabeled PEP, DiR, and PEP + DiR.

Article Snippet: Primary antibodies included CD63 (1:1000, R&D Systems, Minneapolis, MN, USA, #MAB50482), Flotillin (1:1000, Abcam, Waltham, MA, USA, #ab133497), CD9 (1:1000, Cell Signaling Technology, Danvers, MA, USA, #13403S), CD41 (1:1000, NovusBiologicals, Centennial, CO, USA, #NBP1-84581), GAPDH (1:2000, Cell Signaling Technology, 2118S), and actin (1:1000, LI-COR, #926-42212).

Techniques: Membrane, Labeling, Standard Deviation, Transmission Assay, Electron Microscopy, Western Blot, Immunocytochemistry, Control, MANN-WHITNEY, Two Tailed Test

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Microfluidic transection injury and high-shear thrombus formation demonstrate increased hemostatic efficacy of cold-stored platelets and in vitro resuscitation in induced coagulopathy models

doi: 10.3389/fbioe.2025.1568113

Figure Lengend Snippet: Stenotic microfluidic device assessed in vitro hemostatic resuscitation platelet dynamics. (A) Representative images of clot formed in the stenotic channel of the microfluidic device (platelets, CD41, white). (B) Quantified platelet deposition by the fold change in the CD41 mean fluorescent intensity (MFI) is illustrated in healthy controls (WB) and DC and TP samples. (C) Percent occurrence of substantial thrombus formation in the microfluidic device is shown for hemostatic resuscitation experiments in the DC model and (D) corresponding platelet deposition quantification of recipient platelets (CD41 recipient ). (E,G) Percent occurrence of substantial thrombus formation for hemostatic resuscitation experiments in the thrombocytopenia coagulopathy model and (F,H) corresponding platelet deposition quantification of recipient platelets (CD41 recipient ) is shown for RT and CS platelet product mixing conditions. MFI curves are presented as mean ± standard error of the mean.

Article Snippet: For stenotic microfluidic experiments, platelet product aliquots were incubated with Fc receptor block (Biolegend; Human TruStain FcX; 422302; 1:600) and fluorescently tagged CD41 antibody (Janelia Fluor ® 646) (Novus Biologicals; NB100-2614JF646; 1:600).

Techniques: In Vitro

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Microfluidic transection injury and high-shear thrombus formation demonstrate increased hemostatic efficacy of cold-stored platelets and in vitro resuscitation in induced coagulopathy models

doi: 10.3389/fbioe.2025.1568113

Figure Lengend Snippet: RT and CS platelet dynamics in a stenotic microfluidic device. (A) Platelet deposition quantification of recipient platelets (CD41 recipient ) and (B) RT platelet products (CD41 product ). (C) Platelet deposition quantification of recipient platelets (CD41 recipient ) and (D) CS platelet products (CD41 product ). D2, D5, and D7 are shown for both storage conditions. Data are presented as mean ± standard error of the mean.

Article Snippet: For stenotic microfluidic experiments, platelet product aliquots were incubated with Fc receptor block (Biolegend; Human TruStain FcX; 422302; 1:600) and fluorescently tagged CD41 antibody (Janelia Fluor ® 646) (Novus Biologicals; NB100-2614JF646; 1:600).

Techniques:

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Microfluidic transection injury and high-shear thrombus formation demonstrate increased hemostatic efficacy of cold-stored platelets and in vitro resuscitation in induced coagulopathy models

doi: 10.3389/fbioe.2025.1568113

Figure Lengend Snippet: Transected injury microfluidic device assessed blood loss with coagulopathy models to evaluate hemostatic resuscitation. (A) Top–down view schematic representation of the transection injury microfluidic device with assembly, dimensions, and coatings used to assess in vitro hemostatic resuscitation. (B) Representative image of the clot formed in the vascular channel region of the microfluidic device (platelets, CD41, white). (C) Blood loss from healthy controls (WB) and DC and TP samples measured via endpoint microfluidic outflow. Data were analyzed via two paired t-tests comparing paired WB controls within each coagulopathy model. *p < 0.05; **p < 0.01. (D) Hemostatic resuscitation with RT platelet products at D2 were evaluated with mass outflow curves shown and (E) corresponding endpoint microfluidic outflow. Data were analyzed via mixed-effects analysis and Tukey’s post hoc test using a 95% confidence interval. **p < 0.01. All data are presented as mean ± standard deviation or individual values with bars representing mean ± standard deviation.

Article Snippet: For stenotic microfluidic experiments, platelet product aliquots were incubated with Fc receptor block (Biolegend; Human TruStain FcX; 422302; 1:600) and fluorescently tagged CD41 antibody (Janelia Fluor ® 646) (Novus Biologicals; NB100-2614JF646; 1:600).

Techniques: In Vitro, Standard Deviation

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Microfluidic transection injury and high-shear thrombus formation demonstrate increased hemostatic efficacy of cold-stored platelets and in vitro resuscitation in induced coagulopathy models

doi: 10.3389/fbioe.2025.1568113

Figure Lengend Snippet: RT and CS products assessed in the thrombocytopenia model in the transected injury microfluidic device. (A) Representative images from paired healthy control (WB), TP sample, and TP sample mixed with platelet product are shown. TP sample platelets (CD41 recipient , blue) and product platelets (CD41 product , red). (B) Blood loss measured via endpoint microfluidic outflow from hemostatic resuscitation experiments combining storage temperature and age. Data were analyzed via mixed-effects analysis and Tukey’s post hoc test using a 95% confidence interval. *p < 0.05; ***p < 0.001. (C,D) Blood loss segmented by storage temperature. RT product data were analyzed via mixed-effects analysis and Tukey’s post hoc test using a 95% confidence interval. Cold-storage product data were analyzed via ANOVA and Tukey’s post hoc test using a 95% confidence interval. **p < 0.01. (E) Blood loss from TP + RT/CS samples stratified by storage age. (F) Blood loss compared in early-storage (D2) versus late-storage (D5 and D7) products. Early- versus late-storage age data were analyzed via an unpaired t-test. **p < 0.01. All data are presented as mean ± standard deviation or individual values, with bars representing mean ± standard deviation.

Article Snippet: For stenotic microfluidic experiments, platelet product aliquots were incubated with Fc receptor block (Biolegend; Human TruStain FcX; 422302; 1:600) and fluorescently tagged CD41 antibody (Janelia Fluor ® 646) (Novus Biologicals; NB100-2614JF646; 1:600).

Techniques: Control, Standard Deviation

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Microfluidic transection injury and high-shear thrombus formation demonstrate increased hemostatic efficacy of cold-stored platelets and in vitro resuscitation in induced coagulopathy models

doi: 10.3389/fbioe.2025.1568113

Figure Lengend Snippet: Endpoint thrombus composition in the transected injury microfluidic device. (A) Representative image from the TP sample resuscitated with RT platelet products at D2 is shown. TP sample platelets (CD41 recipient , blue) and product platelets (CD41 product , red). (B) CD41 product area quantification and (C) CD41 recipient area quantification are shown highlighting storage at D5 and D7. (D) Calculated ratio of CD41 recipient over CD41 product area is shown for all storage days. ***p < 0.001. All data are presented as individual values, with bars representing mean ± standard deviation.

Article Snippet: For stenotic microfluidic experiments, platelet product aliquots were incubated with Fc receptor block (Biolegend; Human TruStain FcX; 422302; 1:600) and fluorescently tagged CD41 antibody (Janelia Fluor ® 646) (Novus Biologicals; NB100-2614JF646; 1:600).

Techniques: Standard Deviation