Supplementary Figures S1-S26 from The Immunomechanical Checkpoint PYK2 Governs Monocyte-to-Macrophage Differentiation in Pancreatic Cancer

Supplementary Fig. S1 shows representative images of human adjacent normal and PDAC tissues post-surgery, alongside representative images of Masson's trichrome staining for human adjacent normal and PDAC tissues. Supplementary Fig. S2 depicts quality control results of spatial transcriptomics, identifies cell types in human normal pancreas and PDAC tissues, and maps the spatial distribution of major cell types in sections of human adjacent normal and PDAC tissues. Supplementary Fig. S3 displays representative mIHC staining images of human adjacent normal and PDAC tissues. Supplementary Fig. S4 illustrates the mIHC staining and quantification of LYVE1+ macrophages in soft and stiff PDAC tumors. Supplementary Fig. S5 presents the mRNA expression levels of mechanosensitive ion channels and integrins in BM cells cultured on soft, stiff, and TCPS surfaces. Supplementary Fig. S6 illustrates the gating strategy for bone marrow-derived monocytes and macrophages, along with representative monocyte gating in BM cells cultured on soft, stiff, and TCPS surfaces. Supplementary Fig. S7 displays the quality control results of scRNA-seq data derived from human blood and PDAC samples. Supplementary Fig. S8 presents a spatiotemporal analysis of monocytes and macrophages in blood and PDAC tissues from human PDAC patients. Supplementary Fig. S9 illustrates the subcutaneous simulation of PDAC models by injecting stiff or soft hydrogels mixed with KPC cells. Supplementary Fig. S10 presents the protein expression levels of FAK in PYK2-deficient monocytes/macrophages. Supplementary Fig. S11 illustrates the influence of PYK2 on macrophage mechanotransduction in vivo. Supplementary Fig. S12 illustrates the polarization profiles of WT and Ptk2b KO BMDMs under stimulation with LPS, IL-4/IL-13, or conditioned medium. Supplementary Fig. S13 shows PYK2 deficient BMDMs exhibit impaired cell migration. Supplementary Fig. S14 illustrates the impact of PIEZO1 agonists or inhibitors on monocyte differentiation. Supplementary Fig. S15 shows the role of PYK2 as a transcriptional co-regulator in conjunction with AP-1 and/or SP-1. Supplementary Fig. S16 illustrates the effects of PYK2 deficiency in myeloid precursors on the TME and tumor growth in mouse PDAC models. Supplementary Fig. S17 displays the gating strategy and quantitative analysis of myeloid cells and lymphocytes in tissues from mouse PDAC models. Supplementary Fig. S18 presents the gating strategy and quantitative analysis of T cells within a co-culture system of T cells and monocytes/macrophages. Supplementary Fig. S19 illustrates the impact of myeloid-specific PYK2 deficiency on the TME and its enhancement of immunotherapy efficacy in mouse PDAC models. Supplementary Fig. S20 presents the quality control metrics for scRNA-seq data of PDAC tumors from WT + Iso, WT+αPD-1, Ptk2b KO + Iso and Ptk2b KO+αPD-1 groups. Supplementary Fig. S21 depicts the transcriptional profiles of monocytes, macrophages, and fibroblasts across single and combination treatment groups. Supplementary Fig. S22 shows the impact of myeloid-specific Ptk2b deficiency on fibrosis in mouse PDAC tissues. Supplementary Fig. S23 showed the distinct functional profiles of fibroblasts across experimental groups. Supplementary Fig. S24 illustrates the analysis of CD8+ T cells using flow cytometry and mIHC across single and combination treatment groups. Supplementary Fig. S25 shows the palpation-based tumor size measurements for orthotopic PDAC tumors in mice. Supplementary Fig. S26 shows the full Western blot images alongside their corresponding quantitative analyses.