Journal: The Journal of Experimental Medicine

Article Title: KLF family members control expression of genes required for tissue macrophage identities

doi: 10.1084/jem.20240379

Figure Lengend Snippet: Impaired development of LCMs in myeloid-specific KLF2-deficient mice. (A) Flow cytometry of cells in peritoneal cavity lavage from the indicated mice, gated on live, single, CD3 − B220 − CD19 − cells. Numbers adjacent to gates indicate the percentage of total live events for each gate. (B) Bar graphs depicting the percentage of total live cells and total number of cells for the following populations (representative gates shown in A) as measured by flow cytometry: CD11b + F4/80 + , SCM (CD11b + F4/80 + MHCII hi DNAM-1 hi ), LCMs (CD11b + F4/80 + MHCII − DNAM-1 − ), ICAM2 + GATA6 + LCM (CD11b + F4/80 + MHCII − DNAM-1 − ICAM2 + GATA6 + ), and transitional (CD11b + F4/80 + MHCII mid DNAM-1 mid ). (C) Dimensionality reduction via UMAP of combined CD11b + F4/80 + cells from LysM +/+ Gata6 fl/fl and LysM +/+ Klf2 fl/fl (gated together as LysM +/+ controls), LysM Cre/+ Gata6 fl/fl , or LysM Cre/+ Klf2 fl/fl mice, downsampled to normalize cells per genotype. The resulting projection was overlayed with supervised gates for LCMs, transitional, and SCMs as shown in A. The upper panel shows all cells, while the lower panels show only cells from the indicated genotypes. (D) Heatmaps of defining marker expression for SCM, LCM, and transitional cell clusters overlaid onto the UMAP projections from C. Data in A are from one experiment representative of six independent experiments; data in B are combined from six independent experiments, LysM +/+ Gata6 fl/fl ( n = 9), LysM Cre/+ Gata6 fl/fl ( n = 6), LysM +/+ Klf2 fl/fl ( n = 5), LysM Cre/+ Klf2 fl/fl ( n = 7), CX3CR1-Cre − Klf2 fl/fl ( n = 4), and CX3CR1-Cre + Klf2 fl/fl ( n = 5). Significance determined by ordinary two-way ANOVA with multiple comparisons and Šidák’s correction. Asterisks denote: ****P < 0.0001, ***P = 0.0006, **P = 0.0021, and *P = 0.033.

Article Snippet: The following mice were used in this study: C57BL/6J (000664), B6.SJL (B6.Ptprc a Pep c /BoyJ, 002014), JaxBoy (C57BL/6J-Ptprcem6Lutzy/J, 033076), LysM Cre (Lyz2Cretm1[Cre]Ifo, 004781), Klf2 fl/fl (gift from Jerry B. Lingrel, University of Cincinnati, Cincinnati, OH, USA; deceased), LysM Cre Gata6 fl/fl (gift from Paul Kubes, University of Calgary, Calgary, Canada), LysM Cre Klf4 fl/fl (floxed Klf4 allele: MMRRC 29877), CX3CR1-Cre (Tg[Cx3cr1-cre]MW126Gsat/Mmucd, MMRRC: 036395-UCD), CD11c-Cre Klf4 flfl (Tg[Itgax-cre]1-1Reiz/J, 008068, cross generated in-house), GFP-KLF2 (B6[C]-Klf2tm1.1Khog/JmsnJ, gift from Stephen Jameson and Kristin Hogquist labs, University of Minnesota, Minneapolis, MN, USA), and CD45.1 × CD45.2 F1 (generated in our colony by crossing C57BL/6J to B6.SJL [B6.Ptprc]).

Techniques: Flow Cytometry, Marker, Expressing

Journal: bioRxiv

Article Title: Two-Stage CD8 + CAR T-Cell Differentiation in Patients with Large B-Cell Lymphoma

doi: 10.1101/2025.03.05.641715

Figure Lengend Snippet: ( a , b ) UMAP depicting single-cell transcriptomes of peripheral blood CD8 + T cells colored by cell cluster ( a ) or density contours of each cluster ( b ). The exhausted-like EM cluster is located on the lower half of the UMAP. ( c ) Density maps depicting expression levels of major T-cell genes and proteins, divided into categories. In the “receptor” category, proteins are placed directly beneath the corresponding gene. ( d ) Flow plots for validating transcriptomic data. Plots depict expression of CD39 and CD57 (top row) or CX3CR1 (bottom row) in CD8 + CAR T cells at each timepoint from P2. Exhausted-like EM CAR T cells were expected to be T exp -specific with high CD39 and low CD57/CX3CR1 expression. ( e , f ) Heat map depicting normalized expression of major gene sets from Wherry et al. ( e ). Expression of two of the gene sets were depicted as violin plots ( f ), ordered by decreasing expression level per cluster. Expression levels were compared to that of the cluster with highest expression via Wilcoxon Rank-Sum test, with p-values adjusted for multiple hypotheses testing using the Benjamini-Hochberg method, whereby **** indicates p<0.0001, *** indicates p<0.001. ( g ) Volcano plot depicting differentially expressed genes between EM and exhausted-like EM CD8 + CAR T cells. Genes were colored according to direction of upregulation. ( h ) Enrichment plots for select gene sets differentially expressed between EM and exhausted-like EM CD8 + CAR T cells.

Article Snippet: Then, cells were incubated for 30 minutes at 4°C in the dark with a staining solution containing AF647-labeled CD19-tetramers (3 nM final concentration), BV421-labeled anti-CD3ε (clone SK7, BioLegend, 344833), AF488-labeled anti-CD8α (clone SK1, BioLegend, 344716), PE-labeled anti-CX3CR1 (clone 2A9-1, BioLegend, 341603), PE/Cy7-labeled anti-CD39 (clone A1, BioLegend, 328211), and BV605-labeled anti-CD57 (clone QA17A04, BioLegend, 393303).

Techniques: Expressing

Journal: Immunity & Ageing : I & A

Article Title: CD31 + naïve T cells associate with immunosenescence and responsiveness to multiple vaccines in older adults

doi: 10.1186/s12979-025-00504-0

Figure Lengend Snippet: Statistically significant differences in flowsom identified T cell subsets between age groups divided into two sections based on being vaccine response associated or not. Arrows indicate whether the given subset is higher (↑) or lower (↓) in the group1 compared to group2

Article Snippet: For surface antigens CCR4 BV605 (clone: L281H4, supplier: BD Biosciences), CCR6 BV711 (G034E3, Biolegend), CCR7 BV421 (G043H7, Biolegend), CD127 APC-R700 (HIL-7R-M21, BD Biosciences), CD14 Superbright 436 (61D3, ThermoFisher), CD159a APC (REA110, Miltenyi Biotech),, CD19 Superbright 436 (HIB19, ThermoFisher), CD25 cFluorBYG710 (BC96, Cytek), CD27 APC-H7 (M-T271, BD Biosciences), CD28 BV650 (CD28.2, Biolegend), CD3 BUV805 (SK7, BD Biosciences), CD31 BUV563 (L133.1, BD Biosciences), CD38 APC/Fire 810 (HIT2, Biolegend), CD4 cFluorYG584 (SK3, Cytek), CD45RA BUV395 (5H9, BD Biosciences), CD56 Superbright 436 (TULY56, ThermoFisher), CD57 BB515 (NK-1, BD Biosciences), CD8 cFluor V547 (SK1, Cytek), CD95 PE-Cy5 (DX2, Biolegend), CX3CR1 BUV737 (2A9-1, BD Biosciences), CXCR3 PE-Cy7 (G025H7, Biolegend), CXCR5 BV750 (RF8B2, BD Biosciences), HLA-DR BV570 (L243, Biolegend), ICOS BUV661 (DX29, BD Biosciences), KIR2D PE (NKVFS1, Miltenyi Biotech), KIR3DL1/DL2 PE (5.133, Miltenyi Biotech), KRLG1 VioBlue (REA261, Miltenyi Biotech), PD-1 BV785 (EH12.2H7, Biolegend), TCR γδ PerCP-Vio 700 (REA591, Miltenyi Biotech), TIGIT BV480 (741182, BD Biosciences), for intracellular/intranuclear antigens, CTLA-4 PE-CF594 (BNI3, BD Biosciences), Foxp3 BB700 (236 A/E7, BD Biosciences), Helios Alexa Fluor 647 (22F6, Biolegend) were used.

Techniques: Significance Assay