cd74  (Cell Signaling Technology Inc)

Journal: Nature Neuroscience

Article Title: Cerebrovascular vulnerability and fibrosis in human brain aneurysms

doi: 10.1038/s41593-026-02326-9

Figure Lengend Snippet: a , Heatmap with hierarchical clustering showing cell-to-cell differential interaction strength in brain aneurysms when compared to control cerebrovasculature. y -axis, source; x -axis, target. Red, increased interaction strength in aneurysms; blue, decreased interaction strength in aneurysms. EC, endothelial cell; SMC, smooth muscle cell; FbM, fibromyocyte; FB, fibroblast; aFB, activated fibroblast; mFB, myofibroblast; TC, T-cell; NK, natural killer cell; BC, B-cell; pDC, plasmacytoid dendritic cell; cDC, conventional dendritic cell; pvMϕ, perivascular macrophage; Mo, monocyte; aMϕ, APC5 + macrophage; MG, microglia; AC, astrocyte; Neu, neuron; OL, oligodendrocyte; OPC, oligodendrocyte precursor cell. b , Circle plot of differential interaction strength between activated perivascular fibroblasts (aFB) and myeloid cell populations in aneurysms. Red, increased interaction strength in aneurysms; blue, decreased interaction strength in aneurysms. Line thickness proportional to interaction strength. c , Cell-to-cell communication pathways across cell populations ranked on their differences of overall information flow within inferred networks in aneurysms (red) and control (gray). Red text, pathways statistically over-represented in brain aneurysms; gray text, pathways statistically over-represented in controls; black text, not significant. d , Chord plot showing outgoing cell communication pathways originating from myeloid cells to activated perivascular fibroblasts in aneurysms. Arrow thickness is proportional to interaction strength. e , Scatter-plot of pathways ranked by differential incoming ( y -axis) and outgoing ( x -axis) interaction strength in aFB in brain aneurysms relative to controls. Light blue, aneurysm-specific. Bold, top-nominated outgoing pathway emerging from aFB. f , Representative confocal microscopy analysis of immunostaining CD68 + (magenta) macrophages showing colocalization with CD74 receptor (yellow) and MIF ligand (cyan) in unruptured brain aneurysm (top) and control middle cerebral artery (bottom). DAPI (blue) labels cell nuclei. Colocalization, white. Scale bar, 50 µm. g , Representative confocal microscopy analysis of CD68 + macrophages (magenta) and MIF–CD74 proximity interaction (yellow) in unruptured brain aneurysm (top) and control middle cerebral artery (bottom). DAPI (blue) labels cell nuclei. Colocalization, white. Scale bar, 50 µm.

Article Snippet: Next, we added anti-CD74 antibody (MedChemExpress) at concentration of 5 μg ml −1 and incubated for 30 min followed by washing with medium.

Techniques: Control, Confocal Microscopy, Immunostaining

Journal: Nature Neuroscience

Article Title: Cerebrovascular vulnerability and fibrosis in human brain aneurysms

doi: 10.1038/s41593-026-02326-9

Figure Lengend Snippet: a , Violin plot of the established vascular destabilizing genes MMP9 and SPP1 across cell populations. EC, endothelial cell; SMC, smooth muscle cell; FbM, fibromyocyte; FB, fibroblast; aFB, activated fibroblast; mFB, myofibroblast; TC, T-cell; NK, natural killer cell; BC, B-cell; pDC, plasmacytoid dendritic cell; cDC, conventional dendritic cell; pvMϕ, perivascular macrophage; Mo, monocyte; aMϕ, APC5 + macrophage; MG, microglia; AC, astrocyte; Neu, neuron; OL, oligodendrocyte; OPC, oligodendrocyte precursor cell. b , Enriched biological processes from marker genes in ACP5 + macrophages ordered by statistical significance Benjamini-Hochberg adjusted P < 0.05; dashed line, Benjamini-Hochberg adjusted P cutoff of 0.05. c , Bar graph showing quantitative polymerase chain reaction (qPCR) analysis of MMP9 (left) and ACP5 (right) expression in primary human macrophages treated with interferon (IFN), macrophage inhibitory factor (MIF), and CD74 neutralizing antibody (anti-CD74). Mean ± s.e.m. MMP9: Media vs IFN ( P = 0.2479, ns); IFN vs IFN + MIF ( P = 0.0047, **); IFN + MIF vs IFN+anti-CD74 ( P = 0.0046, **); IFN + MIF vs IFN + MIF+anti-CD74 ( P = 0.017, *). ACP5: Media vs IFN ( P = 0.8114, ns); IFN vs IFN + MIF ( P = 0.0052, **); IFN + MIF vs IFN+anti-CD74 ( P = 0.009, **); IFN + MIF vs IFN + MIF+anti-CD74 ( P = 0.009, **). One-way ANOVA with Tukey’s multiple comparisons test (n = 3 samples). d . Boxplot of the distance (µm) from each aMϕ to its nearest neighbor of each listed cell type (n = 770 aMϕ). Dashed line marks 100 µm. Center lines indicate medians; box limits indicate the 25th and 75th percentiles; whiskers extend to the most extreme data points no more than 1.5 × IQR from the box; points beyond the whiskers are shown as outliers.

Article Snippet: Next, we added anti-CD74 antibody (MedChemExpress) at concentration of 5 μg ml −1 and incubated for 30 min followed by washing with medium.

Techniques: Marker, Real-time Polymerase Chain Reaction, Expressing

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Wheat germ agglutinin (WGA), Hematoxylin and eosin (H&E), Masson trichrome, TUNEL, DCF staining, and protein markers for cardiac injury and apoptosis in 8-week-old WT and CD74 −/− mice with or without DOX (5 mg/kg, i.p. once per week for 4 weeks) administration. (A) Representative WGA (upper row), H&E staining (middle row) and Masson staining (bottom row) from respective myocardial sections. (B) Quantitative analysis of cardiomyocyte cross-sectional area in WGA staining. (C) Quantitative analysis of interstitial fibrosis in MASSON staining. (D, E) Quantitative analysis of ANP and BNP level; Insets: Representative gel blots of ANP and BNP using specific antibodies ( β -actin for loading control). (F) TUNEL (upper row) and DCF (lower row) staining of myocardial sections. (G) Quantitative analysis of TUNEL-positive cardiomyocytes. (H) Quantitative analysis of DCF staining; (I–K) Representative gel blots and quantitative analysis of Cleaved-caspase 3, Bcl2 and Bax. (L) Representative DCF of AMCMs from WT and CD74 −/− mice with or without DOX exposure. (M) Pooled fluorescence intensity of DCF in AMCMs. Mean ± SEM; n = 11 images from 3 mice (panels A–C) per group, n = 6 mice (panels D, E, I–K) per group, n = 10–15 images from 3 to 5 mice (panels F–H, L, M), ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: TUNEL Assay, Staining, Control, Fluorescence

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Protection of DOX-induced ferroptosis and cardiotoxicity with pharmacological inhibition of CD74. (A) Timeline of DOX and Amifostine treatments; DOX was administered to C57BL/6J mice for a period of 4 weeks. Beginning in the fifth week, Amifostine (80 mg/kg, once daily) was administered for an additional week. (B) Representative M-mode echocardiographic images. (C–F) Pooled analysis of left ventricular ejection fraction, fractional shortening, normalized LV mass, and heart weight normalized to tibial length (HW/TL). (G, H) Serum IL-6 and TNF- α levels. (I) Representative H&E staining micrographs from respective groups. (J) Quantitative analysis of cardiomyocyte area. (K) Representative Masson trichrome staining images. (L) Quantitative analysis of interstitial fibrosis. (M, N) 4-HNE staining images and 4-HNE positive area. (O) Malondialdehyde (MDA) levels in heart tissues. (P–R) Representative gel blots and quantitative analysis of ANP, BNP using specific antibodies ( β -Actin as loading control); and (S) Schematic diagram illustrating summarized experimental findings and cell signaling pathways involved in CD74 ablation-evoked benefit against DIC. Mean ± SEM; n = 6 mice (panels B–H, P–R) or n = 10 per group from 3 mice (panels I–N) or n = 12 from 3 mice (panel O) per group, ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Inhibition, Staining, Control, Protein-Protein interactions

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Change of CD74 levels in plasma from breast cancer patients with or without DOX exposure, myocardial tissues from mice subject to DOX-induced cardiomyopathy (DOX, 5 mg/kg, i.p. once per week for 4 weeks), and adult mouse cardiomyocytes exposed to DOX (1 μmol/L or 24 h). (A) Volcano plot of CD74 expression in mouse hearts with or without DOX exposure using GSE224157 dataset. (B, C) Immunofluorescence staining of CD74 (green) co-localized with cTnT (red) and CD68 (red) in mouse heart tissues with or without DOX exposure. (D, E) Fluorescence intensity of CD74. (F–K) In the RAW 264.7 macrophage and AMCM cell co-culture system, CD74 was knocked down in RAW 264.7 macrophages to discern its inhibitory effect on cardiomyocyte function with or without DOX exposure. Cell co-culture was performed using Transwell co-culture between RAW 264.7 macrophages and cardiomyocytes; (F) Resting cardiomyocyte cell length; (G) Cardiomyocyte peak shortening; (H) Cardiomyocyte maximal velocity of shortening (+d L /d t ); (I) Maximal velocity of relengthening (–d L /d t ); (J) Cardiomyocyte time-to-peak 90% shortening (TP 90 ); (K) Time-to-90% relengthening (TR 90 ). (L) Representative gel blots of CD74 in DIC from plasma of breast cancer patients, mouse hearts and adult mouse cardiomyocytes (AMCMs) with or without DOX exposure using specific antibodies (Transferrin or β -actin for loading control, transferrin for plasma protein). (M–O) Quantitative analysis of CD74 levels. (P) Association of CD74 with cTnT in plasma from DIC patients. Mean ± SEM; n = 6 images from 3 mice (panels B–E) per group, n = 25 cells (panels F–K), n = 6 (panel G) or 12 patients (panels J) and n = 6 mice (panels H and I) per group, ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Clinical Proteomics, Expressing, Immunofluorescence, Staining, Fluorescence, Co-Culture Assay, Control

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Echocardiographic, cardiomyocyte contractile and intracellular Ca 2+ properties in 8-week-old WT and CD74 −/− mice with or without DOX (5 mg/kg, i.p. once per week for 4 weeks) exposure. (A) Timeline of DOX treatments. (B) Representative echocardiographic images. (C) Left ventricular end-diastolic diameter (LV EDD). (D) Left ventricular end-systolic diameter (LV ESD). (E) Septal wall thickness. (F) Posterior wall thickness in diastole (PWT, d). (G) Ejection fraction. (H) Fractional shortening. (I) Body weight. (J) Normalized LV mass (to body weight). (K) Heart rate. (L) Resting cardiomyocyte cell length. (M) Cardiomyocyte peak shortening. (N) Cardiomyocyte maximal velocity of shortening (+d L /d t ). (O) Maximal velocity of relengthening (–d L /d t ); (P) Cardiomyocyte time-to-peak 90% shortening (TP 90 ). (Q) Time-to-90% relengthening (TR 90 ). (R) Baseline Fura-2 fluorescence intensity (FFI). (S) Electrically stimulated rise in FFI (ΔFFI) and (T) Intracellular Ca 2+ decay rate. Mean ± SEM; n = 13 mice per group (panels B–J), n = 48 (panels L–Q) or 32 (panels R–T) per group from 3 to 5 mice, ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Fluorescence

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Mitochondrial ultrastructure and function, and Western blot analysis of autophagy in myocardium from 8-week-old WT and CD74 −/− mice with or without DOX (5 mg/kg, i.p. once weekly for 4 weeks) insult. (A) Transmission electron microscopy (TEM) images showing mitochondria ultrastructure. (B) Individual mitochondrial area (μm 2 ). (C) Mitochondrial circularity (width-to-length ratio). (D) Representative JC-1 micrographs for aggregates (top row), monomer (middle row) and merged (bottom row) fluorescence. (E) Pooled JC-1 analysis. (F–I) OCR curves and quantification of basal respiration, maximal respiration, and ATP production in AMCMs from WT and CD74 −/− mice with or without DOX challenge. (J–L) Mitochondrial respiration (complex I/III, complex II/III, and complex IV) in AMCMs from WT and CD74 −/− mice with or without DOX challenge. (M) Representative gel blots of UCP2, LC3B and p62 using specific antibodies ( α -Tubulin or GAPDH as loading control). (N–P) Quantitative analysis of UCP2, LC3B and p62 levels. Mean ± SEM; n = 10 visual fields (panels B and C) or n = 15 images (panels D, E) from 5 to 6 mice, and n = 8 (panels F–I) per group or n = 6 (panels J–L) per group from 3 mice, and n = 6 mice per group (panels M–P), ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Western Blot, Transmission Assay, Electron Microscopy, Fluorescence, Control

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: CD74/RRM2 interaction, its impact on CD74/RRM2 distribution and ferroptosis in 8-week-old WT and CD74 −/− mice with or without DOX (5 mg/kg, i.p. once per week for 4 weeks) exposure. (A) The protein–protein interaction network was conducted using the STRING database. (B) Representative gel blots depicting RRM2 and p53 levels (GAPDH or NKATPase as loading control, NKATPase for cell membrane protein). (C, D) Quantitative analysis of RRM2 in cytoplasm and cell membrane. (E) Quantitative analysis of p53 level. (F) KEGG enrichment from transcriptomics data of DIC mice ( GSE224157 ). (G) Application of gene set variation analysis (GSVA) for calculation of the activity level of ferroptosis pathway from transcriptomics data of DIC mice ( GSE224157 ). (H) Intracellular Fe 2+ detected using the FerroOrange probe in AMCMs from WT and CD74 −/− mice with or without DOX exposure. (I) Fluorescence intensity of intracellular Fe 2+ . (J) Representative gel blots of GPX4, SCL7A11 and NCOA4 ( β -Actin or Vinculin as loading control). (K–M) Quantitative analysis of GPX4, SCL7A11 and NCOA4 levels. Mean ± SEM; n = 6 mice (panels B–E, J–M) or n = 10 fields from 3 mice (panels H, I) per group, ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Control, Membrane, Transcriptomics, Activity Assay, Fluorescence

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Interaction of CD74 with RRM2 through 180–215 amino acid domain for recruitment of RRM2 onto cytomembrane. (A, B) Structure-based interface analysis between CD74 and RRM2. CD74–RRM2 complex structure with interaction hot-spot residues labeled using PRISM. (C) Graph illustrating construction of CD74 (total length) and mutants (Delta 180–215 aa, Delta 220–250 aa, Delta 180–250 aa). (D) IP analysis of CD74 tagged with Flag (CD74-Flag) and RRM2 tagged with His (RRM2-His) from AMCMs from WT and CD74 −/− mice with or without DOX exposure (5 mg/kg, i.p. once per week for 4 weeks). (E) Representative fluorescent images for co-localization of CD74 (green) with RRM2 (red) using H9C2 cells. (F, G) IP analysis of CD74 tagged with Flag (CD74-Flag) and RRM2 tagged with His (RRM2-His); H9C2 cells were divided into 4 groups (CD74-Flag, Delta 180–215 aa, Delta 220–250 aa, Delta 180–250 aa) prior to transfection of RRM2 using plasmid DNA, Confocal image exhibiting H9C2 cells transfected with indicated plasmids, stained with Dil (blue), anti-CD74 (green), and anti-RRM2 (red) antibodies. (H–K) Fluorescence intensity curve of CD74 and RRM2 in co-localization images.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Labeling, Transfection, Plasmid Preparation, Staining, Fluorescence

Journal: Acta Pharmaceutica Sinica. B

Article Title: CD74 deficiency protects against doxorubicin cardiotoxicity through RRM2-mediated regulation of ferroptosis

doi: 10.1016/j.apsb.2026.01.028

Figure Lengend Snippet: Protection of DOX-induced ferroptosis and cardiotoxicity with pharmacological inhibition of CD74. (A) Surface plasmon resonance (SPR) assay the binding of CD74 and DOX. (B) Molecular docking simulation of the binding between Amifostine and CD74. (C–I) AMCMs were exposed to DOX (1 μmol/L) for 24 h, prior to an additional 24-h treatment with or without Amifostine (10 μmol/L), Pifithrin- β (10 μmol/L), Kevetrin (80 μmol/L), LIP-1 (200 nmol/L), or GW8510 (4 μmol/L). (C) Cell length. (D) Peak shortening. (E) Maximal velocity of shortening (+d L /d t ). (F) Maximal velocity of relengthening (– d L /d t ). (G) Time-to-peak 90% shortening (TP 90 ). (H) Time-to-90% relengthening (TR 90 ). (I) Quantified BODIPY fluorescence intensity. (J) Representative BODIPY staining images from respective cell groups. Mean ± SEM; n = 26 (panels C–H) or 20 (panels I, J) per group from 3 mice, ∗ P < 0.05 between indicated groups.

Article Snippet: For CD74 inhibition using Amifostine (CAS No.: 20537-88-6, MCE), control and DIC mice were treated with Amifostine (80 mg/kg, dissolved in 10% DMSO and 90% saline, i.p.) once daily for 1 week .

Techniques: Inhibition, SPR Assay, Binding Assay, Fluorescence, Staining

Journal: Neuro-Oncology

Article Title: MIF-CD74 signaling drives immune modulation in medulloblastoma

doi: 10.1093/neuonc/noag020

Figure Lengend Snippet: Cell-to-cell communications within the TIME for human Group3 and Group4 MB. (A) The UMAP plot of cell type clusters show the identification of 15 distinct cell clusters from the single-cell RNA-seq data of Group3 MB samples. OPC: Oligodendrocyte Precursor Cell. APC: Astrocyte Precursor Cell. M2_1: M2 macrophages cluster 1. M2_2: M2 macrophages cluster 2. NSC: Neural stem cell. Complement-M, complement macrophage. inflam dendritic cell: inflammatory dendritic cell. (B) The violin plot of the marker gene expression for each cell cluster. (C) CellCrossTalker predicted ligand-receptor-mediated tumor-immune cell communications using scRNA-seq data from Group3 MB samples. The vertical axis represents ligands and their corresponding receptors, while the horizontal axis indicates the sender cell type associated with the ligand and the receiver cell type associated with the receptor. (D) CellCrossTalker predicted ligand-receptor-mediated communications between M2 macrophages and tumor cells, M2 macrophages and T cells, M2 macrophages and B cells, as well as M2 macrophages and myeloid cells, using scRNA-seq data from Group3 MB samples. The vertical axis represents ligands and their corresponding receptors, while the horizontal axis shows the sender cell type associated with the ligand and the receiver cell type associated with the receptor. (E) The UMAP plot of cell type clusters show the identification of 13 distinct cell clusters. Single-cell RNA-seq data from Group4 MB samples were obtained from Hendrikse et al. (F) The heatmap illustrates the expression of marker genes used to annotate each cell cluster. (G) CellCrossTalker predicted ligand-receptor-mediated cell-cell communications using scRNA-seq data from Group4 MB samples. The vertical axis represents ligands and their corresponding receptors, while the horizontal axis shows the sender cell type associated with the ligand and the receiver cell type associated with the receptor. (H) The heatmap shows the interaction strength of MIF-CD74 between various pairs of cell types, as predicted by CellCrossTalker using scRNA-seq data from Group4 MB samples. (I) CellCrossTalker predicted ligand-receptor-mediated communication co-receptors between tumor cells and immune compartment, using scRNA-seq data from Group3 and Group4 MB samples. The vertical axis represents ligands and their corresponding receptors and co-receptors, while the horizontal axis shows the sender cell type associated with the ligand and the receiver cell type associated with the receptor.

Article Snippet: In brief, tissue sections were incubated in Tris-EDTA buffer (cell conditioning 1; CC1) at 95 ̊C for 1-h to retrieve antigenicity, followed by incubation with CD74 antibody (Origene CF507339 ) at 1:500 for 1-h.

Techniques: Single Cell, RNA Sequencing, Marker, Gene Expression, Expressing

Journal: Neuro-Oncology

Article Title: MIF-CD74 signaling drives immune modulation in medulloblastoma

doi: 10.1093/neuonc/noag020

Figure Lengend Snippet: Restricted expression of CD74 and MIF in human normal tissues and MB subgroups. (A) Graph depicting the expression of CD74 (ENSG00000019582.14) in normal human tissue RNA-sequencing data obtained from the GTEx consortium. The dataset comprises 7859 samples across 31 distinct normal tissues, with sample sizes ranging from 5 to 1152 samples per tissue. Expression levels are presented as relative expression levels in transcripts per million (TPM). (B) CD74 protein expression in normal human cerebellum. Scale bar, 100 um. (C) Graph depicting the expression of MIF (ENSG00000240972.1) in normal human tissue RNA-sequencing data obtained from the GTEx consortium. The dataset comprises 7859 samples across 31 distinct normal tissues, with sample sizes ranging from 5 to 1152 samples per tissue. Expression levels are presented as relative expression levels in transcripts per million (TPM). (D) MIF protein expression in normal human cerebellum. Scale bar, 100 um. (E-J) Protein levels across human MB subgroups and subtypes (based on DNA methylome classification) from Ayrault cohort for the three selected proteins: CD74, CD68 and MIF. Boxplots show median (line), upper and lower quartiles (boxes), and lines extending to highest and lowest observations (whiskers). (K) CD74 immunohistochemistry staining analysis of paired human pediatric diagnostic (left) and relapse (right) MB samples. Black arrows depict CD74 positivity. Scale bar represents 100 µM. (L) Immunohistochemistry membrane staining depicting CD74 expression across a subgrouped human diagnostic MB tissue microarray. Scale bar represents 200 µM.

Article Snippet: In brief, tissue sections were incubated in Tris-EDTA buffer (cell conditioning 1; CC1) at 95 ̊C for 1-h to retrieve antigenicity, followed by incubation with CD74 antibody (Origene CF507339 ) at 1:500 for 1-h.

Techniques: Expressing, RNA Sequencing, Immunohistochemistry, Staining, Diagnostic Assay, Membrane, Microarray

Journal: Neuro-Oncology

Article Title: MIF-CD74 signaling drives immune modulation in medulloblastoma

doi: 10.1093/neuonc/noag020

Figure Lengend Snippet: Inhibition of CD74 via the lateral ventricle demonstrates immune-modulation and decreased tumor burden in primary and relapsed MB. (A) IHC of treatment-naïve primary (top row) and radiation-treated (bottom row) GTML allografts displaying H&E, CD74, and MIF expression within the tumor. Results representative of 3 independent replicates. Much of the tumor outside the cerebellum was removed for downstream RNA sequencing analyses, Scale bar, 100 µm. (B) Experimental schematic for evaluating the CD74-MIF blocking peptide C36L1. GTML primary or relapse cells expressing firefly luciferase were allografted into the cerebellum of FVBNRJ mice. C36L1 was administered either intraperitoneally or locoregionally through the lateral ventricle at time of engraftment and 1-week after, C36L1 vehicle was used as a control. Bioluminescence imaging was conducted to monitor tumor engraftment, progression, and/or regression up to 14 days post-therapy. At the endpoint, the tumor and CNS were harvested and assessed for immune infiltration and tumor burden. (C) Growth rate of mice bearing primary GTML allografts ( n = 3-4 per group) treated with vehicle control (black), C36L1 peptide through the lateral ventricle (red), or C36L1 peptide intraperitoneally (blue) was determined by calculating the slope of tumor growth between day 7 and 14 (endpoint). Line represents mean growth rate with individual points representing individual mice. Points below the dashed line indicate tumor regression. Statistical significance was calculated by two-way ANOVA with Tukey’s post-test. (D) UMAP projection of flow cytometry analysis of tumor/cerebellum of mice treated vehicle control or the C36L1 peptide via the lateral ventricle. The left panel display cells colored by experimental group (Control - black, C36L1 delivered via lateral ventricle - red, C36L1 delivered intraperitoneally - blue), highlighting the distribution and intensity target expression across different cell populations within the tumor and cerebellum of treated and control mice. The visualization provides insights into the immune cell infiltration and its association with the treatment groups. (E) UMAP projection of flow cytometry results, colored by the expression of CD74. (F) UMAP projection of flow cytometry results, colored by the expression of MHCII. (G) UMAP projection of flow cytometry results, colored by the expression of CD11b. (H) IHC of control (top row) and peptide-treated (bottom row) GTML primary tumor allografts displaying H&E, CD3, F4/80 and CD74 within the tumor. Results representative of 3 independent replicates, Scale bar, 100 um. (I) Growth rate of mice bearing recurrent GTML allografts ( n = 5 per group) treated with vehicle control (black) or C36L1 peptide through the lateral ventricle (blue) was calculated by calculating the slope of tumor growth between day 7 and 14 (endpoint). Line represents mean growth rate with individual points representing individual mice. Points below the dashed line indicate tumor regression. Statistical significance was calculated by two-way ANOVA with Tukey’s post-test. (J) Bar chart to illustrate the proportion of tumor-associated immune cells identified as microglia. A higher proportion of microglia is observed in the TME of CD36L1 peptide treatment versus scrambled control. (K) Bar chart to illustrate the level of CD74 expression in the TME of mice treated with CD36L1 versus scrambled control. (L) Bar charts displaying the proportion of CD38+ cells in the microglia population. Statistical analysis was performed using a two-way ANOVA with Tukey’s post-test to compare the groups. Error bars represent the SD. Significant differences between groups are indicated by * P < .05, ** P < .01, and *** P < .001.

Article Snippet: In brief, tissue sections were incubated in Tris-EDTA buffer (cell conditioning 1; CC1) at 95 ̊C for 1-h to retrieve antigenicity, followed by incubation with CD74 antibody (Origene CF507339 ) at 1:500 for 1-h.

Techniques: Inhibition, Expressing, RNA Sequencing, Blocking Assay, Luciferase, Control, Imaging, Flow Cytometry

Journal: Therapeutic Advances in Medical Oncology

Article Title: S100A4 characterize antigen-presenting cancer-associated fibroblasts and predicts surgical outcomes in relapsed ovarian cancer

doi: 10.1177/17588359261436959

Figure Lengend Snippet: Immune spatial interactions and prognostic significance of CD74 + S100A4 + antigen-presenting CAFs in ROC. (a, b) Spatial proximity analysis between CAF subpopulations and CD4 + T cells using mIHC and computational phenotyping. (a) Representative mIHC images showing spatial relationships between αSMA + , S100A4 + , CD74 + S100A4 + CAFs, and CD4 + T cells. Lines indicate nearest neighbor distances between cells. Scale bar, 50 µm. (b) Boxplot quantification of mean number of CD4 + T cells within 20 µm radius of each CAF subtype. CD74 + S100A4 + CAFs displayed significantly closer proximity to CD4 + T cells. ( p < 0.05) as shown in representative image (a). (c–e) Differences in CD74 + S100A4 + CAFs distribution and their spatial relationship with CD4 + T cells between patients achieving R0 versus Non-R0. (c) Representative images of mIHC staining illustrating differences in spatial cell arrangement. Scale bar, 200 µm. (d) Quantification of CD74 + S100A4 + CAFs densities (cells/mm²) and (e) mean count of CD4 + T cells within 20 µm of CD74 + S100A4 + CAFs between R0 and Non-R0 groups. (f, g) Prognostic significance of S100A4 + apCAFs based on multi-dataset transcriptomic analysis. (f) Forest plot showing HR of S100A4 + apCAFs-associated gene signature across 11 ovarian cancer datasets. Each horizontal black square represents the HR estimate from an individual dataset, and the horizontal line indicates the 95% CI. The overall HR for S100A4 + apCAFs is shown at the bottom, with the dashed vertical line indicating the reference value HR = 1. (g) In the TCGA ovarian cancer cohort, patients were stratified into a high-expression group (top 30%, n = 68, shown in blue) and a low-expression group (bottom 30%, n = 68, shown in red) based on the expression levels of the top 100 S100A4 + apCAFs signature genes. The Kaplan–Meier survival curves compare overall survival between these groups. The x -axis represents time since diagnosis (in months), and the y -axis indicates overall survival probability. CAF, cancer-associated fibroblasts; CI, confidence interval; HR, hazard ratio; mIHC, multiplex immunohistochemistry; ROC, relapsed ovarian cancer; S100A4, S100 calcium-binding protein A4; TCGA, The Cancer Genome Atlas; αSMA, α-smooth muscle actin.

Article Snippet: Sections underwent antigen retrieval in citrate or EDTA buffer, followed by endogenous peroxidase blocking with 3% H 2 O 2 , serum blocking, and overnight incubation at 4°C with primary antibodies: αSMA (#19245, Cell Signaling Technology, Danvers, MA, USA), FAP (#ab207178, Abcam, Cambridge, UK), S100A4 (#13018, Cell Signaling Technology, Danvers, MA, USA), PDPN (#26981, Cell Signaling Technology, Danvers, MA, USA), PAX8 (#1F8-3A8, Thermo Fisher Scientific, Waltham, MA, USA), and CD74 (#77274, Cell Signaling Technology, Danvers, MA, USA).

Techniques: Staining, Expressing, Biomarker Discovery, Multiplex Assay, Immunohistochemistry, Binding Assay