Journal: Cell Reports Medicine

Article Title: Multimodal immune phenotyping reveals microbial-T cell interactions that shape pancreatic cancer

doi: 10.1016/j.xcrm.2024.101397

Figure Lengend Snippet: T cell and microbial co-localization define distinct cellular neighborhoods in human pancreatic cancer (A) Study design for (B)–(D) (n = 185 for cold nests and n = 77 for hot nests pooled from 16 specimens). (B) Images of CD8 (yellow), CD68 (brown), tumor epithelium (teal, CK19), FOXP3 (purple), and nuclei (blue, hematoxylin). Scale bars, 1 mm (top) and 50 μm (bottom). (C) Quantification of CD8 + (left), FOXP3 + (middle), and CD68 + (right) cells. (D) Ratio of CD8 + cells to FOXP3 + cells. (E) Study design for (F) and (H)–(J). (F) Principal-component analysis (n = 5 for hot and cold tumor nests from 1 specimen). (G) 16S rRNA levels (n = 14 for cold and hot tumor nests from 3 patient specimens). (H and I) Heatmap (H) and enrichment score (I) for the relative expression of genes associated with response to bacterium (GO: 0009617) in hot and cold stroma. (J) Volcano plot of DEGs. (K) Multiplex immunohistochemistry images of CD8 (yellow), PIGR (purple), and nuclei (blue, hematoxylin). (L) Quantification of PIGR (n = 49 for cold nests and n = 46 for hot nests pooled from 7 specimens). (M) Multiplex immunohistochemistry images of CD8 (teal), CD20 (yellow), CD74 (purple), and nuclei (blue, hematoxylin). (N) Quantification of CD74 (n = 71 for cold nests and n = 45 for hot nests pooled from 10 specimens). (O) Summary of cell markers and proteins. For (K) and (M), scale bars, 2 mm (left) and 50 μm (right), and dashed lines indicate tumor epithelium. Statistical significance was calculated using a two-tailed Mann-Whitney test. Data are represented as violin plots (center line, median; top and bottom lines, upper and lower quartiles) and scatterplots (mean + SD or mean). PDAC, pancreatic ductal adenocarcinoma.

Article Snippet: Mouse monoclonal anti-human CD74 (LN2) , Abcam , ab9514; RRID:AB_2075504.

Techniques: Expressing, Multiplex Assay, Immunohistochemistry, Two Tailed Test, MANN-WHITNEY

Journal: Cell Reports Medicine

Article Title: Multimodal immune phenotyping reveals microbial-T cell interactions that shape pancreatic cancer

doi: 10.1016/j.xcrm.2024.101397

Figure Lengend Snippet: T cell infiltration into tumors occurs independent of the gut and tumor microbiome (A) Study design for (B)–(K). (B) 16S rRNA levels in tumor and stool from mice orthotopically injected with cold (69) tumor cells (n = 10) or hot (2838c3) tumor cells and treated with (n = 15) and without (n = 20) antibiotics. (C and D) Quantification of CD3 + (C) and CD8 + and FOXP3 + (D) T cells from hot tumors of mice treated with (n = 19) or without (n = 13) antibiotics. (E) 16S rRNA levels in tumor from mice orthotopically injected with cold (69) tumor cells and treated with (n = 5) and without (n = 5) antibiotics. (F) Quantification of CD8 + and FOXP3 + T cells from cold tumors of mice treated with (n = 5) and without (n = 5) antibiotics. (G) Number of DEGs. (H) Bar graph displaying overrepresentation analysis of DEGs in indicated gene sets. (I and J) Tumor weights at day 20. (K) Quantification of intra-tumoral CD19 + cells. (L and M) Mean fluorescence intensity (MFI) of MHC class II (L) and CD206 (M) on CD11b + F4/80 + intra-tumoral macrophages. Data were pooled from two to three experiments (B–D and J–M) or are representative of two independent experiments (E–I). Statistical significance was calculated using one-way ANOVA with Dunnett’s test (B, L, and M) and a two-tailed Mann-Whitney test (C–F and I–K). Data are represented as scatterplots (mean ± SD). NS, not significant.

Article Snippet: Mouse monoclonal anti-human CD74 (LN2) , Abcam , ab9514; RRID:AB_2075504.

Techniques: Injection, Fluorescence, Two Tailed Test, MANN-WHITNEY

Journal: Cell Reports Medicine

Article Title: Multimodal immune phenotyping reveals microbial-T cell interactions that shape pancreatic cancer

doi: 10.1016/j.xcrm.2024.101397

Figure Lengend Snippet:

Article Snippet: Mouse monoclonal anti-human CD74 (LN2) , Abcam , ab9514; RRID:AB_2075504.

Techniques: Recombinant, Sequencing, Software

Journal: Cancers

Article Title: HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer

doi: 10.3390/cancers13246179

Figure Lengend Snippet: LN-2 and UMAb231 antibody specificity. ( A ) Western blot analysis of CD74 expression was performed by loading whole cell extracts of MDA-MB-231 control cells (control), MDA-MD-231 cells with CD74 siRNA knockdown (+CCD74 siRNA), and IFNγ stimulated MDA-MD-231 cells (+IFNγ). CD74 protein was detected at 33/35 kDa using either anti-CD74 LN-2 (left-hand panel) or UMAb231 monoclonal antibody (right-hand panel), and at 41/43 kDa with UMAb231 in overexpressing cells. *, **, and *** mark the positions of three independent bands of unknown identity. No signal was detected in siRNA-mediated CD74 knockdown cells (+CD74 siRNA). IHC analysis of FFPE sections of MDA-MB-231 breast cancer control cells compared to ( B ) CD74 siRNA knockdown, and ( C ) IFNγ stimulation showed similar staining patterns with LN-2 and UMAb231 antibodies. Insets in subpanels 1b and 1c show higher magnification images illustrating punctuated stainings of CD74. ( D ) Quantification of stainings showed that IHC immunoreactivity correlated with expression of cellular CD74 protein. Scale bars = 20 µm. The uncropped blot is shown in .

Article Snippet: Recently, a new anti-CD74 mouse monoclonal antibody, UMAb231, was developed by Origene Technologies (Rockville, MD, USA).

Techniques: Western Blot, Expressing, Staining

Journal: Cancers

Article Title: HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer

doi: 10.3390/cancers13246179

Figure Lengend Snippet: IHC staining patterns of UMAb231. Immunoreactivity for CD74 was seen primarily in stromal components, ( A ) in peritumoral fibroblasts, ( B ) endothelial cells, and ( C ) lymphocytes and macrophages, respectively (black arrows). Staining of serial sections with vimentin, CD4, CD8, CD68 or CD31 was used to confirm cell identity. In some samples, CD74 was also expressed in ( D ) tumor cells (black arrow indicates tumor cells), with essentially cytoplasmic and ( E ) membrane staining (black arrow indicates membrane staining). ( F ) Comparative semi-quantitative analysis of the UMAb231 and LN-2 antibodies, showed similar, but not always identical, patterns between the two antibodies. A case is highlighted with no CD74 immunoreactivity in tumor cells with UMAb231 (sub-panel b), but weak to moderate reactivity with LN-2 (sub-panel a, black arrow). In both cases lymphocytes and macrophages stained strongly with either antibody (sub-panels a and b, red arrows, respectively). Scale bars are 100 µm in ( A , D , F ), 25 µm in B, 250 µm in ( C ), and 50 µm in ( E ), respectively.

Article Snippet: Recently, a new anti-CD74 mouse monoclonal antibody, UMAb231, was developed by Origene Technologies (Rockville, MD, USA).

Techniques: Immunohistochemistry, Staining

Journal: Cancers

Article Title: HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer

doi: 10.3390/cancers13246179

Figure Lengend Snippet: IHC analysis of YTMA49. TMA cores were scored in a categorical manner according to CD74 immunoreactivity levels in tumor cells only, as follows: 0 (absent), 1+ (weak), 2+ (moderate) or 3+ (strong) (( A , C , E , G ), respectively). An H-score value was also derived by computer-assisted whole tissue core analysis ( B , D , F , H ). Negative, weak, moderate and strong immunostaining thresholds are yellow, orange, and red, respectively. Intensity scores were correlated to ( I ) TN status and ( J ) node status. Scale bars = 250 µm.

Article Snippet: Recently, a new anti-CD74 mouse monoclonal antibody, UMAb231, was developed by Origene Technologies (Rockville, MD, USA).

Techniques: Derivative Assay, Immunostaining

Journal: Cancers

Article Title: HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer

doi: 10.3390/cancers13246179

Figure Lengend Snippet: Low/high score association with pathology scores, nodal status and TN status (categorical variables).

Article Snippet: Recently, a new anti-CD74 mouse monoclonal antibody, UMAb231, was developed by Origene Technologies (Rockville, MD, USA).

Techniques:

Journal: Cancers

Article Title: HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer

doi: 10.3390/cancers13246179

Figure Lengend Snippet: Survival analysis of breast cancer samples as a function of CD74 expression. ( A ) The prognostic value of CD74 expression was examined in the YTMA49 cohort for all cases, TN cases, and non-TN cases. ( B ) Kaplan–Meier plot showing OS among breast cancer patients according to CD74 expression in a multivariate model correcting for immune cell infiltration in a multivariable Cox proportional hazard model. Correlation between CD74 gene expression and infiltration of immune cells was examined in the breast cancer samples available from TCGA database with the TIMER analysis tool ( http://timer.cistrome.org ) (accessed on 8 March 2019) ( C ) Correlation between CD74 and PD-L1 in 1017 breast carcinomas from TCGA after correcting for tumor-infiltrating immune cells.

Article Snippet: Recently, a new anti-CD74 mouse monoclonal antibody, UMAb231, was developed by Origene Technologies (Rockville, MD, USA).

Techniques: Expressing