Journal: Non-coding RNA Research

Article Title: CircSMAD4 shapes matrix-remodeling TAMs in lung adenocarcinoma

doi: 10.1016/j.ncrna.2026.03.003

Figure Lengend Snippet: circSMAD4 drives tumor-educated M2-like polarization of macrophages and promotes tumor-cell aggressiveness. (A) Workflow for generating TC-hMDMs and TC-BMDMs, circSMAD4 knockdown, and downstream functional assays. (B) RT–qPCR analysis of M1-associated markers (MHC-II [HLA-DRA in TC-hMDMs; H2-Ab1 in TC-BMDMs], NOS2, and CD86) and M2-associated markers (CD163, CD206, and ARG1) in TC-hMDMs and TC-BMDMs. (C) Representative flow-cytometry histograms for HLA-DR, iNOS, CD86, CD163, CD206, and ARG1 in TC-hMDMs. Gating strategy and marker thresholds were defined based on FMO controls (see ). (D) Flow-cytometry quantification of marker-positive cells in TC-hMDMs and TC-BMDMs. (E) ELISA of IL-10, TGF-β, and iNOS in culture supernatants. (F) CCK-8 assays of A549 and LLC cells. (G) Colony-formation assays of A549 and LLC cells with quantification. (H) Bioluminescence-based growth readouts of patient-derived LUAD organoids (PDO #1 and PDO #2) after co-culture with TC-hMDMs. (I) Immunoblot analysis of EMT-related proteins (E-cadherin, N-cadherin, Vimentin) in A549 and LLC cells. (J) Transwell migration and invasion assays of A549 and LLC cells with quantification. Scale bar, 50 μm. ∗P < 0.05; ∗∗P < 0.01; ∗∗∗P < 0.001; ∗∗∗∗P < 0.0001; ns, not significant.

Article Snippet: Human IL-10 was quantified using IL-10 DuoSet ELISA (R&D Systems, Cat# DY217B), and human TGF-β1 was measured using the Human TGF beta-1 ELISA Kit (Invitrogen, Cat# BMS249-4).

Techniques: Knockdown, Functional Assay, Quantitative RT-PCR, Flow Cytometry, Marker, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Derivative Assay, Co-Culture Assay, Western Blot, Migration

Journal: Non-coding RNA Research

Article Title: CircSMAD4 shapes matrix-remodeling TAMs in lung adenocarcinoma

doi: 10.1016/j.ncrna.2026.03.003

Figure Lengend Snippet: circSMAD4 drives tumor-educated M2-like polarization of macrophages and promotes tumor-cell aggressiveness. (A) Workflow for generating TC-hMDMs and TC-BMDMs, circSMAD4 knockdown, and downstream functional assays. (B) RT–qPCR analysis of M1-associated markers (MHC-II [HLA-DRA in TC-hMDMs; H2-Ab1 in TC-BMDMs], NOS2, and CD86) and M2-associated markers (CD163, CD206, and ARG1) in TC-hMDMs and TC-BMDMs. (C) Representative flow-cytometry histograms for HLA-DR, iNOS, CD86, CD163, CD206, and ARG1 in TC-hMDMs. Gating strategy and marker thresholds were defined based on FMO controls (see ). (D) Flow-cytometry quantification of marker-positive cells in TC-hMDMs and TC-BMDMs. (E) ELISA of IL-10, TGF-β, and iNOS in culture supernatants. (F) CCK-8 assays of A549 and LLC cells. (G) Colony-formation assays of A549 and LLC cells with quantification. (H) Bioluminescence-based growth readouts of patient-derived LUAD organoids (PDO #1 and PDO #2) after co-culture with TC-hMDMs. (I) Immunoblot analysis of EMT-related proteins (E-cadherin, N-cadherin, Vimentin) in A549 and LLC cells. (J) Transwell migration and invasion assays of A549 and LLC cells with quantification. Scale bar, 50 μm. ∗P < 0.05; ∗∗P < 0.01; ∗∗∗P < 0.001; ∗∗∗∗P < 0.0001; ns, not significant.

Article Snippet: Human IL-10 was quantified using IL-10 DuoSet ELISA (R&D Systems, Cat# DY217B), and human TGF-β1 was measured using the Human TGF beta-1 ELISA Kit (Invitrogen, Cat# BMS249-4).

Techniques: Knockdown, Functional Assay, Quantitative RT-PCR, Flow Cytometry, Marker, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Derivative Assay, Co-Culture Assay, Western Blot, Migration

Journal: bioRxiv

Article Title: Linking live-cell behavior to transcriptional responses across perturbations using dynamic caging

doi: 10.64898/2026.05.05.723043

Figure Lengend Snippet: A, Schematic of Treg experimental design. Briefly, individual Tregs in CCEs were activated with T cell TransAct and stimulated with various cytokine cocktails for 6 days (Conditions I-IV) or 13 days (Conditions I+III). Tregs stimulated for 13 days were also stained for surface IL-10 protein levels. B , Uniform manifold approximation and projection (UMAP) of batch-corrected transcriptomic profiles of all 21,922 T cells from both 6 and 13 day time points colored by (left to right, top to bottom) FOXP3 expression, CellTypist predicted cell types from Hao et al. PBMC atlas (see methods) , experimental stimulation condition, and Leiden cluster. C , Composition plot of experimental condition by Leiden cluster for clusters 1-3. D , Dot plot of Treg activation (red) and suppression (blue) markers for Leiden clusters 1-3. E , Violin plot of tissue Treg scores of cells profiled at day 6 in each experimental condition (see Methods). F , Kernel density estimate plot for cells profiled at day 6 of cell area, and cell cycling score (see Methods). Dotted line represents delineation between low area and high area cells in G. G , Volcano plot of differential expression (DE) analysis between large Tregs (cell area > 3000 pixels, i.e. diameter > 15.4 μm), and small Tregs (cell area < 3000 pixels). Red points: genes upregulated in large cells (right, FDR < 0.05, log2FC > 1), or genes upregulated in small cells (left, FDR < 0.05, log2FC < -1). Shown right are two brightfield images of cells typifying these classes. H , Kernel density estimate (KDE) plots of IL-10 immunofluorescence (IF) signal for cells profiled at day 13 in majority Treg clusters, and cells in majority Tconv. clusters . The dotted line represents delineation between low IL-10 and high IL-10 cells in I. I , Volcano plot of DE analysis between Tconv. cells with high IL10 IF signal (IL-10 IF > 10) and low IL-10 IF signal (IL-10 IF < 10). Red points: genes upregulated in IL-10 IF-high Tconvs. (right, FDR < 0.05, log2FC > 1). Shown right IF images of two cells typifying these classes. J , DE analysis between T cells profiled on day 13 with high eccentricity (eccentricity > 0.9) and cells with low eccentricity (eccentricity < 0.9). Red points: genes upregulated in highly eccentric cells (right, FDR < 0.05, log2FC > 0.5), or genes upregulated in low eccentricity cells (left, FDR < 0.05, log2FC < -0.5). Shown right are two IF images of cells typifying these classes. *Padj<0.05, **Padj<0.01, ***Padj<0.001 .

Article Snippet: On day 13, IL-10 secretion was assessed using the Miltenyi IL-10 Secretion Assay (Cat# 130-090-434).

Techniques: Staining, Expressing, Activation Assay, Quantitative Proteomics, Immunofluorescence

Journal: bioRxiv

Article Title: Linking live-cell behavior to transcriptional responses across perturbations using dynamic caging

doi: 10.64898/2026.05.05.723043

Figure Lengend Snippet: A , Confidence scores of CellTypist predictions for each CD4 T class included in Hao et al. PBMC atlas (all cells included). B , T cell receptor (TCR), and inflammatory T reg signature scores for Leiden clusters 1-3 in (see methods). C , UMAP of 17,967 T cells profiled at 6 days colored by (left to right, top to bottom) experimental condition, CellTypist predicted cell type, tissue Treg score, cell size, and cycling score. D , UMAP of 3,955 T cells profiled at 13 days colored by (left to right) CellTypist predicted cell type, and IL-10 IF signal. E, UMAP of Hao et al. CD4 cells colored by (left to right) cell type, and IL10 expression. F, Dot plot of gene expression across cell types in Hao CD4 T cell atlas. G , Tr1 and Th2 signature scores for IL-10 IF high (IL-10 IF > 10) and IL-10 IF low (IL-10 IF < 10) T conventional cells (see methods). H , Inferred transcription factor activity score for IL-10 IF high and low Tconv cells. Transcription factor inference was performed using a univariate linear model from decoupler and the CollecTRI transcription factor database. I , UMAP of cells profiled at 13 days colored by (left to right) eccentricity, and Leiden cluster. J, Dot plot of gene expression for high (eccentricity > 0.5) and low eccentricity (eccentricity < 0.5) cells in each Leiden cluster in H. P values for gene score comparison and DE analysis were calculated using a two-sided Wilcoxon rank sum test. *Padj<0.05, **Padj<0.01, ***Padj<0.001 .

Article Snippet: On day 13, IL-10 secretion was assessed using the Miltenyi IL-10 Secretion Assay (Cat# 130-090-434).

Techniques: Expressing, Gene Expression, Activity Assay, Comparison