in house tg2 monoclonal antibody dh2  (R&D Systems)

Journal: Nature Communications

Article Title: MALT1 protease inhibition restrains glioblastoma progression by reversing tumor-associated macrophage-dependent immunosuppression in mice

doi: 10.1038/s41467-026-76572-7

Figure Lengend Snippet: a Induction of MALT1 protease activity within macrophages was measured using RAW264.7 cells stably expressing a cleavage-activated luciferase reporter, which emits luminescence upon activation of MALT1 protease or b induction of NF-kB activity within macrophages was measured in RAW-Blue macrophages co-cultured with GL261-GBM cells. Results are shown as fold change relative to control macrophages cultured alone. c Cytokines and chemokines in the cell culture supernatant were measured by cytometric bead array 48 h after co-culturing primary mouse macrophages with GL261-GBM cells in the absence or presence of MALT1i. Cytokine levels are presented as a heat map indicating fold change relative to the control macrophages alone. *Significant difference compared to macrophage control alone. #Significant difference compared to macrophage + GBM co-culture. d , e Changes in macrophage M1/M2-like polarization markers after co-culture with GBM cells. d RAW264.7 mouse macrophages were co-cultured with GL261 mouse GBM cells in the absence or presence of MALT1i or e CARD9i. f NF-kB activity in RAW-Blue macrophages co-cultured with GL261-GBM cells. g Changes in macrophage M2-like polarization markers after co-culture with GBM cells. RAW264.7 mouse macrophages were co-cultured with GL261 mouse GBM cells in the absence or presence of a blocking antibody against mouse Dectin-1 receptor (1–5 µg/mL) or the appropriate Rat IgG2a Isotype Control Antibody (5 µg/mL). h – j The Incucyte live-cell analysis system was used to measure tumor cell apoptosis in real-time. Apoptotic tumor cells are indicated by the overlap (or colocalization) of red and green to produce yellow. h Fluorescent images of RAW264.7 macrophages co-cultured with GBM-GL261 tumor cells ± MALT1i. Scale bar = 400 µM. i Co-cultures of mouse RAW264.7 macrophages with GL261-GBM cells were performed to compare macrophage-dependent tumor cell apoptosis in the absence/presence of MALT1i. j Controls demonstrate that MALT1i has no effect on apoptosis of GL261-GBM tumor cells or RAW264.7 macrophages when either is cultured alone. All values are represented as mean ± SD. Data were analyzed by 1-way ANOVA, followed by Tukey’s multiple-comparisons. P values are indicated in the figure.

Article Snippet: In experiments with the MALT1 inhibitor, macrophages were exposed to MLT-748 (5 μM, catalog no.: HY-115466, Med Chem Express), BRD5529 (50 μM, catalog no.: HY-115497), antibody against murine Dectin-1 (clone R1-8g7) (1–5 μg/mL, catalog no.: mabg-mdect-2, Invivogen) or the appropriate Rat IgG2a Isotype Control Antibody (catalog no.: mabg2a-ctlrt, Invivogen) for 2 h prior to addition of tumor cells and during the entire co-culture time.

Techniques: Activity Assay, Stable Transfection, Expressing, Luciferase, Activation Assay, Cell Culture, Control, Co-Culture Assay, Blocking Assay, Cell Analysis