Journal: Cancers

Article Title: Multiplex Imaging Mass Cytometry Reveals Prognostic Immunosuppressive Subpopulations and Macrophage-Driven Metastasis in Osteosarcoma

doi: 10.3390/cancers17172780

Figure Lengend Snippet: Imaging mass cytometry (IMC) and cell phenotyping analysis of OS tissue microarray (TMA). ( A ) Representative IMC images displaying nuclei, single-marker ( left ), and double-marker ( right ) staining of cells within the OS tumor microenvironment (TME). The left panel highlights the most abundant cell populations, and the right panel shows immunosuppressive cells (IMSC) with respective markers used in this study. Representative examples of double-marker-stained cells are shown with a white arrow. ( B ) Box plot illustrating the number of objects (cells) with positive staining for 25 single markers ( x -axis) that passed quality control (QC) across 51 TMA cores ( y -axis). The y -axis is presented on a log 10 scale. ( C ) Box plot showing the percentage of various detected TME cell types following cell phenotyping across 51 tumor cores. The y -axis is presented on a log10 scale. ( D ) Stacked bar plot depicting the percentage of total detected immune and stromal cells per tumor core. Immune cells include CD68+ (macrophages), CD11b + CD14 + CD15 − (mMDSC), CD11b + CD14 − CD15 + (pMDSC), CD14 + CD16 − CD68 − (cMono), CD14 + CD16 + CD68 − (ncMono), CD15 + CD16 + (neutrophils), CD11c + CD3 − CD14 − CD56 − (dendritic cells, DC), CD56 + CD16 + (cNK), CD56 + GZMB + (aNK), CD56 + CD3 + (tNK), CD3 + CD4 + (helper T cells), CD3 + CD8 + (cytotoxic T cells), and CD3 + FoxP3 + (Treg). Stromal cells include CD31 + (endothelial cells), Pan CK + (pan-epithelial-like cells), E-cadherin + (epithelial-like cells), and aSMA + (fibroblasts).

Article Snippet: Stained TMAs were imaged using the Hyperion Imaging Mass Cytometry (IMC) system (Standard BioTools, South San Francisco, CA, USA).

Techniques: Imaging, Mass Cytometry, Microarray, Marker, Staining, Control

Journal: Cancers

Article Title: Multiplex Imaging Mass Cytometry Reveals Prognostic Immunosuppressive Subpopulations and Macrophage-Driven Metastasis in Osteosarcoma

doi: 10.3390/cancers17172780

Figure Lengend Snippet: The effect of M2 macrophages on pulmonary metastasis in OS. ( A ) Upper panel: Cell morphology of THP-1-derived M0, M1, and M2 macrophages taken at 20× magnification with a light microscope. Small, round THP-1 cells treated with 10 ng/mL PMA differentiated into large, elongated M0 macrophages. M0 cells were further treated with 20 ng/mL of IFN-γ and 10 pg/mL of LPS or 20 ng/mL of IL-4 and IL-13 to induce M1 and M2 macrophage differentiation, respectively, resulting in larger, stellate-shaped cells with increased cytoplasmic-to-nuclear volume. Lower panel: Flow cytometry analysis of CD14, HLA-DR (M1 marker), and CD163 (M2 marker) expression levels in differentiated or polarized macrophages. ( B ) Confirmation of THP-1 differentiation and polarization of M0 into M1 and M2 using canonical markers specific for macrophage subtypes. Upper panel: Flow cytometry analysis of marker expression. Lower panel: ELISA analysis of conditioned media collected after 72 h of rest in macrophage maintenance medium (MMM). ( C ) ( Left ): Representative H&E-stained images of metastatic nodules in resected lungs from orthotopic xenograft mouse models intratibially injected with 143B cells alone, 143B with M0 macrophages (M0 CC), or 143B with M2 macrophages (M2 CC). Metastatic nodules are indicated with yellow arrows. ( Right ): Comparisons of primary tumor volumes calculated as 0.5 × height × width 2 (upper) and the number of metastatic nodules in mouse models with different cell injections after 5 weeks. Each dot represents an individual mouse. Error bars indicate standard deviations, and asterisks denote statistical significance (* p < 0.05; **** p < 0.0001; unmarked = not significant).

Article Snippet: Stained TMAs were imaged using the Hyperion Imaging Mass Cytometry (IMC) system (Standard BioTools, South San Francisco, CA, USA).

Techniques: Derivative Assay, Light Microscopy, Flow Cytometry, Marker, Expressing, Enzyme-linked Immunosorbent Assay, Staining, Injection

Journal: The Journal of Pathology

Article Title: Characteristic miRNA profiles represent clinicopathological diversity of small cell lung cancer

doi: 10.1002/path.6458

Figure Lengend Snippet: Immune cell profiling in SCLC samples with high and low miR‐9‐5p expression. (A) Representative H&E staining, IMC signals, tissue classification via machine learning, and cell annotation in SCLC samples with high and low miR‐9‐5p expression. (B) Box plot illustrating the proportion of stromal area in miR‐9‐5p‐high SCLC (red) and miR‐9‐5p‐low SCLC (green). The top and bottom ends of the box represent the 75th and 25th percentiles, and the line in the middle of the box indicates the median. (C) Heatmap showing the proportion of each cell component in SCLC samples in the intra‐tumoral and stromal areas. (D) Box plots illustrating the proportion of CD8 + T cells and CD163 − macrophages in the intra‐tumoral area and plasma cells in the stromal area. The top and bottom ends of the box represent the 75th and 25th percentiles, and the line in the middle of the box indicates the median. (E) Representative images of CD8 + T cells, CD163 + /CD163 − macrophages, and plasma cells in miR‐9‐5p‐high SCLC. * p < 0.05. IMC, imaging mass cytometry.

Article Snippet: In brief, FFPE tissue sections were stained with a cocktail of metal‐conjugated antibodies using the Human Maxpar Immuno‐Oncology Imaging Mass Cytometry Panel Kit (#201508; Fluidigm, South San Francisco, CA, USA), together with 141Pr‐CD38 (#3141018D; Fluidigm), 143Nd‐Vimentin (#3143027D; Fluidigm), 147Sm‐CD163 (#3147021D; Fluidigm), and 151Eu‐CD31 (#3151025D; Fluidigm).

Techniques: Expressing, Staining, Clinical Proteomics, Imaging, Mass Cytometry