Journal: PLOS Pathogens

Article Title: Regulatory T cell-like response to SARS-CoV-2 in Jamaican fruit bats ( Artibeus jamaicensis ) transduced with human ACE2

doi: 10.1371/journal.ppat.1011728

Figure Lengend Snippet: Four bats were transduced with human ACE2 and infected 5 days later. After 12 days, splenocytes were recovered for AIM testing and gene expression analsysis. A. Splenocytes from hACE2-transduced bats 12 days post infection with SARS-CoV were cultured with or without SARS-CoV-2 nucleocapsid peptide library for 6 hours in the presence of anti-CD40 blocking antibody (AIM testing) or 24 hours without anti-CD40 blocking antibody (cellular RNA). B. Cells were stained with anti-CD154 and anti-CD4 and analyzed by flow cytometry. Violin plots for the individual bats, plus the pooled sample and rank data. In the presence of peptide, CD154 expression was significantly increased on splenocytes (red) compared to no peptide controls (blue) (Wilcoxon rank sum test). C. Heat map of splenocyte qPCR (ΔΔCq) gene expression in response to peptide stimulation determined 3 of the 4 bats expressed a profile consistent with a regulatory T cell response. D. Peptide-induced gene expression changes in splenocyte cultures of bats 1, 2 and 4 from panel C; IL-10, TGFβ and CD4 gene expression were significantly elevated, whereas CXCR4 was nearly significant (two-tailed t-test).

Article Snippet: Test samples were then resuspended in a 1:20 dilution of anti-mouse CD40 PE (Tonbo, clone FGK45), or anti-human CD154 PE (Tonbo, clone 5C8) in FACS buffer and incubated for 30 minutes at 4°C in the dark.

Techniques: Transduction, Infection, Gene Expression, Cell Culture, Blocking Assay, Staining, Flow Cytometry, Expressing, Two Tailed Test

Journal: PLoS Medicine

Article Title: Inhibition of Endothelial Activation: A New Way to Treat Cerebral Malaria?

doi: 10.1371/journal.pmed.0020245

Figure Lengend Snippet: HBEC-5i were left unstimulated or were activated with LMP-420 alone, TNF alone, TNF with LMP-420, LT alone, or LT with LMP-420. After an overnight incubation, cells were detached and analyzed by flow cytometry after immunolabeling for ICAM-1, VCAM-1, and CD40. Results are expressed as dotplots of mean fluorescence intensity of surface expression of the antigens measured in six experiments (Dunn's test, ** corresponds to p < 0.01).

Article Snippet: HBEC-5i were then harvested and labeled by indirect labeling using mouse anti-human CD54 (ICAM-1 [84H10]) and CD106 (VCAM-1 [1G1]) antibodies (Beckman-Coulter Immunotech, Marseille, France), CD40 monoclonal antibody (mAb) (B-B20, Diaclone, Besançon, France), and CD36 mAb (FA6–152, gift from L. Edelman, Institut Pasteur Paris) as the first step.

Techniques: Incubation, Flow Cytometry, Immunolabeling, Fluorescence, Expressing

Journal: BMC Neuroscience

Article Title: Mir-199a-3p aggravates neuroinflammation in an Alzheimer’s disease transgenic mouse model by promoting M1-polarization microglia Mir-199a-3p 通过促进 M1 极化小胶质细胞加重阿尔茨海默病转基因小鼠模型中的神经炎症

doi: 10.1186/s12868-025-00965-5

Figure Lengend Snippet: MiR-199a-3p promoted M1 polarization in BV2. A BV2 were transfected with miR-199a-3p inhibitor or negative control or mimic. QRT-PCR was used to evaluate miR-199a-3p levels in BV2 24 h after transfection. B Western blotting analysis of iNOS, IL-1β and Arg-1 protein levels in BV2 after miR-199a–3p inhibition and overexpression. C–E Relative expression levels of each protein. F The proportion of M1/M2 microglia is approximately equal to the ratio of iNOS to Arg-1. G Transfected with the miR-199a-3p inhibitor, negative control, or mimics simulated with or without LPS, BV2 was harvested to stain for flow cytometry analysis. Gates correspond to resting microglia (CD11b+/CD45+), M1 type microglia (CD40+/CD206-) and M2 type microglia (CD40-/CD206+). H , I Proportion of CD40 + and CD206 + cells of the whole sample. J The proportion of M1/M2 microglia is approximately equal to the ratio of CD40+/CD206- to CD40-/CD206+

Article Snippet: Prior to antibody staining, cell receptors were blocked with anti-murine CD16/CD32 Fc receptor (Elabscience Biotechnology Co., Ltd, China) at 4 °C for 10 min.After blocking, BV2 cells were double-stained with PerCP-conjugated anti-murine CD40 (Elabscience Biotechnology Co., Ltd, China) and PE-conjugated anti-murine CD206 (Elabscience Biotechnology Co., Ltd, China) for 20 min.Cells were analyzed using a CytoFLEX flow cytometer (Beckman Coulter Biotechnology, USA).Ten thousand events were recorded, with M1-type microglia identified by positive expression of CD40 and M2-type microglia identified by positive expression of CD206.Furthermore, flow cytometry analysis of microglia in mouse hippocampal tissue was performed according to the following protocol.After euthanasia, part of the hippocampal tissue was dissected and placed in a 6-well plate.The tissue was homogenized using a filter head, and 2 mL of PBS was added to prepare a cell suspension by passing the solution through a 40 μm cell strainer.

Techniques: Transfection, Negative Control, Quantitative RT-PCR, Western Blot, Inhibition, Over Expression, Expressing, Staining, Flow Cytometry