Journal: bioRxiv

Article Title: Cell-type specific innate immune responses shape rabies virus tropism

doi: 10.1101/2021.07.26.453802

Figure Lengend Snippet: (A) Schematic overview of the genomic organization of Tha-eGFP and Th2P-4M-eGFP. Tha-eGFP and Th2P-4M-eGFP harbour the same genetic background except for two mutations introduced in viral P-protein (W265G, W287V) and four mutations introduced in the M protein (R77K, D100A, A104S, M110L) ( , , ). For imaging purposes, the eGFP sequence was introduced after the M protein gene sequence as described previously . (B) Representative immunofluorescence pictures of SK-N-SH, SVGp12, and HMC3 cells upon infection with Tha-eGFP and Th2P-4M-eGFP. (C) Quantification of eGFP + cells in human CNS monocultures upon Tha-eGFP or Th2P-4M-eGFP infection. (D) Quantification of eGFP + cells in human CNS monocultures upon Tha-eGFP or Th2P-4M-eGFP infection and subsequent IFN-α treatment at 24 hours post-infection. (B-D) Cells were infected with Tha-eGFP or Th2P-4M-eGFP (MOI 0.5) and imaged at 48 hours post-infection. All experiments were performed three times (n=3) independently. (C-D) Each dot represents imaging of one well of a 96-well-plate (approx. 8×10 3 cells/well). Bars show mean ± SD with a Tukey’s multiple comparisons test (**** adjusted p-value<0.0001, *** adjusted p-value<0.001, ** adjusted p-value<0.01, * adjusted p-value<0.05). eGFP = Enhanced Green Fluorescent Protein.

Article Snippet: The human neuroblastoma cell line SK-N-SH (ATCC ® HTB-11 TM ), the human astrocyte-like cell line SVGp12 (ATCC® CRL8621TM), and the human microglial cell line HMC3 (ATCC ® CRL-3304 TM ) were cultured in Dulbecco’s Modified Eagle Medium (10566016, Invitrogen) supplemented with 10% heat-inactivated fetal bovine serum (FCS, S182H-500, Eurobio) at 37°C and 5% CO 2 .

Techniques: Imaging, Sequencing, Immunofluorescence, Infection

Journal: bioRxiv

Article Title: Cell-type specific innate immune responses shape rabies virus tropism

doi: 10.1101/2021.07.26.453802

Figure Lengend Snippet: Astrocyte-like SVGp12 and microglia-like HMC3 protect neuroblastoma cells from Tha-eGFP infection in vitro . (A) Quantification of eGFP + cells in mono- and co-cultures. (B) Quantification of TUBB3 + eGFP + cells in mono- and co-cultures . (A – B) Mono- or co-cultures were infected with Tha-eGFP or Th2P-4M-eGFP (MOI 0.5) and imaged at 48 hours post-infection using the OPERA Phenix ® High Content Screening System (Perkin Elmer). (C-E) Quantification of eGFP + cells in SK-N-SH monocultures. After infection with Tha-eGFP or Th2P-4M-eGFP (MOI 0.5), SK-N-SH cells were incubated with the filtered fresh medium (FM) or filtered conditioned medium (CM) from non-infected or homotypic infected (MOI 5, 24 hours post-infection) SK-N-SH (C), SVGp12 (D), or HMC3 cells (E). In detail, THA-eGFP-infected cells were only treated with supernatants from cells previously infected with THA-eGFP. Correspondingly, TH2P-4M-eGFP-infected cells were only treated with supernatants from cells previously infected with TH2P-4M-eGFP. Cells were imaged at 48 hours post-infection. (A – E) All experiments were performed three times (n=3) independently. Each dot represents imaging of one well of a 96-well-plate (approx. 8×10 3 cells/well). Bars show mean ± SD with a Tukey’s multiple comparisons test (**** adjusted p-value<0.0001, *** adjusted p-value<0.001, ** adjusted p-value<0.01, * adjusted p-value<0.05). CM = conditioned medium; eGFP = Enhanced Green Fluorescent Protein; FM = filtered fresh medium; TUBB3 = Class III Beta-Tubulin.

Article Snippet: The human neuroblastoma cell line SK-N-SH (ATCC ® HTB-11 TM ), the human astrocyte-like cell line SVGp12 (ATCC® CRL8621TM), and the human microglial cell line HMC3 (ATCC ® CRL-3304 TM ) were cultured in Dulbecco’s Modified Eagle Medium (10566016, Invitrogen) supplemented with 10% heat-inactivated fetal bovine serum (FCS, S182H-500, Eurobio) at 37°C and 5% CO 2 .

Techniques: Infection, In Vitro, High Content Screening, Incubation, Imaging

Journal: bioRxiv

Article Title: Cell-type specific innate immune responses shape rabies virus tropism

doi: 10.1101/2021.07.26.453802

Figure Lengend Snippet: (A) Intracellular protein concentrations of SK-N-SH cells upon Tha or Th2P-4M infection. (B) Intracellular protein concentrations of hiNeurons upon Tha or Th2P-4M infection. (C) Intracellular protein concentrations of astrocyte-like SVGp12 cells upon Tha or Th2P-4M infection. (D) Intracellular protein concentrations of fetal pAstrocytes upon Tha or Th2P-4M infection. (E) Intracellular protein concentrations of microglia-like HMC3 cells upon Tha or Th2P-4M infection. (A-E) Cells were infected with Tha or Th2P-4M (MOI 5) and intracellular protein concentration was quantified at 48 hours post-infection using the DropArray system (Curiox). All experiments were performed three times (n=3) independently. All bars show mean ± SD with a one-way ANOVA analysis. To correct for multiple testing, the p-value was corrected accordingly (* adjusted p-value<0.0055).

Article Snippet: The human neuroblastoma cell line SK-N-SH (ATCC ® HTB-11 TM ), the human astrocyte-like cell line SVGp12 (ATCC® CRL8621TM), and the human microglial cell line HMC3 (ATCC ® CRL-3304 TM ) were cultured in Dulbecco’s Modified Eagle Medium (10566016, Invitrogen) supplemented with 10% heat-inactivated fetal bovine serum (FCS, S182H-500, Eurobio) at 37°C and 5% CO 2 .

Techniques: Infection, Protein Concentration

Journal: Acta pharmacologica Sinica

Article Title: Prebiotic diet normalizes aberrant immune and behavioral phenotypes in a mouse model of autism spectrum disorder.

doi: 10.1038/s41401-024-01268-x

Figure Lengend Snippet: Fig. 3 Prebiotic diet resolves phagocytic microglia activation in CRB of in utero VPA-exposed mice. Neuroinflammation markers in different brain regions. a Representation from mPFC in the control group stained with anti-Iba1, b number of Iba1+ cells in mPFC, c mean volume of Iba1+ cells in mPFC, d occupancy index of Iba+ cells in mPFC, PBS control: n = 3 mice; VPA control: n = 6 mice; PBS GOS/FOS: n = 6 mice; VPA GOS/FOS: n = 9 mice. e TMEM119 protein expression relative to GAPDH in CRB, and f CD68 protein expression relative to GAPDH in CRB. Analyzed blots were processed in parallel. PBS control: n = 6 mice; VPA control: n = 6 mice; PBS GOS/FOS: n = 5 mice; VPA GOS/FOS: n = 7 mice. *P < 0.05.

Article Snippet: The following primary antibodies were used: TMEM119 (1:1000, Proteintech, 66948-1; marker of microglial activation) [42], CD68 (1:500, Bio-Rad, MCA341R; marker of activated phagocytic microglia) [43], and GAPDH (1:2000, Cell Signaling, 2118) as loading control. iDisco and light sheet imaging The number of serotonergic neurons and the morphological profile of microglia in mouse brains were determined by immunolabeling-enabled three-dimensional imaging of solventcleared organs (iDISCO) [44, 45].

Techniques: Activation Assay, In Utero, Control, Staining, Expressing

Journal: SLAS technology

Article Title: High-throughput cytokine detection platform for evaluation of chemical induced microglial activation

doi: 10.1016/j.slast.2025.100347

Figure Lengend Snippet: (A) A layout map depicting the mouse cytokine array with 62 specified targets. (B) Cytokine profiling from BV2 cell culture medium treated with 100 ng/ml LPS or water (control) for 24 h. (C) Imaging quantification of cytokine array spots of BV2 samples, showing visible cytokines shown in B. The red and blue boxes denoted the position of TNF-α and IL-6, respectively. All values are represented by the mean ± SD ( n = 3 replicates). Statistical analysis was performed using a one-way ANOVA followed by Tukey’s multiple-comparison test, with p -values presented in each graph.

Article Snippet: The murine BV2 microglia cells were purchased from the AcceGen (Fairfield, NJ, USA) and maintained at 90 % RPMI-1640 (ATCC, catalog: 30-2001) supplemented with 10 % FBS as well as 100 U/mL penicillin and 100 μg/mL streptomycin.

Techniques: Cell Culture, Control, Imaging, Comparison

Journal: Acta neuropathologica

Article Title: Anti-pyroglutamate-3 Aβ immunotherapy engages microglia and inhibits amyloid accumulation in transgenic mouse models of Aβ amyloidosis.

doi: 10.1007/s00401-025-02892-5

Figure Lengend Snippet: Fig. 8 Microglia dynamics around individual plaques after antibody treatment in two-photon microscopy study. a Representative in vivo multiphoton microscopy images of Aβ plaques (cyan) and microglia surrounding plaques (purple) at baseline, 2 days and 14 days after injection of ABBV-916 chi or negative control antibody. Some of the plaques (red arrows) had a robust increase of microglia in response to antibody treatment while some of the plaques (blue arrows) did not have increase of microglia. Scale bars = 20 µm. b, c Quantifica- tion of plaque-associated microglia at 2 day and 14 days, respectively, after administration of antibodies. Every dot represents an individual plaque in each mouse. The dotted line represents the averaged micro- glia volume of the negative control group. Blue shadowed area indi- cates the range of mean ± 2×SD of the control group. d, e Compari- son between the ABBV-916 chi and control antibody treated animals at 2 day and 14 days after injection. Data was expressed as mean ± SEM. *p<0.05; **p<0.01, t-test, n=7–8/group

Article Snippet: Images of Aβ plaques, CAA, microglia and vessels were obtained using an Olympus FVMPE-RS Multiphoton Laser Scanning Microscope and an Olympus 25 x dipping objective (NA = 1.05).

Techniques: Microscopy, In Vivo, Injection, Negative Control, Control

Journal: Brain : a journal of neurology

Article Title: Microglial phenotypes in the human epileptic temporal lobe

doi: 10.1093/brain/awy276

Figure Lengend Snippet: (A) changes in form of an amoeboid microglia from the CA1 region and (B) a ramified microglia of the subiculum induced by 2 mM ADP. Two-photon imaging of cells stained with a fluorescent tomato lectin. At left, a microglia imaged at t=0 and towards the right 3D reconstructions from t=0, 5, 12, 30 min after ADP stimulus onset. Scale bar 10μm. Light blue in 3D images, differences between successive reconstructions, shows ruffling of amoeboid and ramified cells. Yellow arrows for ramified microglia point to process retraction between 0 and 5, and between 5 and 12 min. (C) Time course of changes in mean somatic area (circles), arborization area (diamonds) and bleb area (triangles) for amoeboid microglia (n=7, 3 patients) and for (D) ramified microglia (n=7, 2 patients). (E) A longer delay from ADP application to membrane ruffling for ramified than for amoeboid microglia (2-tailed t test, t = -4,554 -, p< 0.001, n=14, 5 patients). (F) Correlation between cellular morphological index and the latency to ruffling. (Pearson correlation, r = 0.9; p=9.0e-6). Amoeboid microglia open circles, ramified microglia filled circles.

Article Snippet: Imaging microglia in tissue slices Living microglia were labelled with tomato lectin conjugated to a fluorescent marker (Dy-light 594, Vector Labs; Schwendele et al., 2012 ).

Techniques: Imaging, Staining