Journal: Advanced Science

Article Title: Bone Marrow Macrophage STING Signaling is a Direct Nexus Linking Marrow Adipose Expansion to NAD + Depletion and Osteogenic Decline in Skeletal Aging

doi: 10.1002/advs.77571

Figure Lengend Snippet: BMAd accumulation and sDPP4 release during skeletal aging activate BMM STING signaling. (a) Heatmap of dysregulated factors (≥1.5‐fold change) in the bone marrow supernatant of 3‐/18‐month‐old male rats identified through 4D label‐free technology ( n = 3 biological replicates). (b) Venn diagram of dysregulated factors in the bone marrow of rats. (c) ELISA of sDPP4 levels in conditioned medium collected from BMAds, Pre‐Ocs, BMMs, Ocs, BMSCs, Obs, and Neu ( n = 6 biological replicates; * p < 0.05 compared to BMAds‐CM). (d) Representative immunofluorescence staining showing that DPP4 was markedly detected in BMAds (Perilipin) in femora from 3‐/24‐month‐old male mice (scale bar: 100 µm). (e) ELISA of sDPP4 levels in conditioned medium collected in 3 M‐/18 M‐BMAds ( n = 6 biological replicates). (f,g) Representative micro‐CT images (f) and quantitative analysis of BV/TV, Tb. N, Tb.Th, and Tb.Sp (g) from PBS‐treated or rDPP4‐treated male mice ( n = 6 biological replicates). (h,i) Representative micro‐CT images (h) and quantitative analysis of BV/TV, Tb. N, Tb.Th, and Tb.Sp (i) from AAV9‐BMAd‐shNC‐treated or AAV9‐BMAd‐sh Dpp4 ‐treated male mice ( n = 6 biological replicates). (j) Representative immunoblots of cGAS, STING, TBK1, IRF3, NF‐κB, and IκB protein/phosphorylation levels in BMMs from PBS‐treated or rDPP4‐treated male mice ( n = 3 biological replicates). (k) Representative immunoblots of cGAS and STING protein/phosphorylation levels in BMMs treated with either rDPP4 alone or in combination with 18 M‐SN ( n = 3 biological replicates). Data are shown as mean ± SD.

Article Snippet: Conversely, rDPP4 (Sino Biological, 10688‐H08H) administration involved once‐weekly intramedullary injections (0.2 μg per injection) for 4 weeks.

Techniques: Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Micro-CT, Analysis, Western Blot, Phospho-proteomics

Journal: Advanced Science

Article Title: Bone Marrow Macrophage STING Signaling is a Direct Nexus Linking Marrow Adipose Expansion to NAD + Depletion and Osteogenic Decline in Skeletal Aging

doi: 10.1002/advs.77571

Figure Lengend Snippet: BMAd‐derived sDPP4 activates BMM STING signaling through TLR4 binding and mtDNA cooperation. (a) Schematic diagram of sDPP4‐mediated STING activation. (b) Representative immunoblots of TLR2 and TLR4 protein levels in BMMs from rDPP4‐treated or SN‐rDPP4‐treated male mice ( n = 3 biological replicates). (c) Representative immunoblots of TAK1 and STING protein/phosphorylation levels in BMMs stimulated with SN‐rDPP4 and treated with C29 or Resatorvid ( n = 3 biological replicates). (d) Molecular docking analysis of the direct binding mode between the TLR4/MD‐2 complex and the DPP4 structure. (e) SPR analysis of the direct interaction between rDPP4 and rTLR4. (f) Co‐Immunoprecipitation analysis of Flag‐ Dpp4 and HA‐ Tlr4 binding. (g) Co‐Immunoprecipitation analysis of DPP4 deletion mutants binding to full‐length TLR4. (h) qPCR analysis of cytosolic DNA extracted from the cytosolic extracts of 3 M‐/24 M‐BMMs ( n = 6 biological replicates). (i,j) Representative immunofluorescence staining of s anti‐dsDNA (green), anti‐Hsp60 (red), and DAPI (blue) in 3 M‐/18 M BMMs (scale bar, 1 µm) (i). The percentage of dsDNA (green) outside of mitochondria (red) was quantified (j) ( n = 15 from 3 biological replicates). (k) Representative immunoblots of cGAS, STING, TBK1, IRF3, NF‐κB, and IκB protein/phosphorylation levels in BMMs treated with EtBr ( n = 3 biological replicates). (l) Flow cytometry analysis and quantification of mtROS production in WT‐3 M‐/WT‐18 M‐BMMs and Sting ΔMφ ‐18 M‐BMMs ( n = 6 biological replicates). (m) Flow cytometry analysis and quantification of mtROS production in SN‐rDPP4‐treated or co‐cultured BMMs with or without Cgas and Sting knockdown ( n = 6 biological replicates). Data are shown as mean ± SD.

Article Snippet: Conversely, rDPP4 (Sino Biological, 10688‐H08H) administration involved once‐weekly intramedullary injections (0.2 μg per injection) for 4 weeks.

Techniques: Derivative Assay, Binding Assay, Activation Assay, Western Blot, Phospho-proteomics, Docking Assay, Analysis, Co-Immunoprecipitation Assay, Immunofluorescence, Staining, Flow Cytometry, Cell Culture, Knockdown

Journal: Advanced Science

Article Title: Bone Marrow Macrophage STING Signaling is a Direct Nexus Linking Marrow Adipose Expansion to NAD + Depletion and Osteogenic Decline in Skeletal Aging

doi: 10.1002/advs.77571

Figure Lengend Snippet: STING signaling promotes NAD + decline in BMMs during skeletal aging by enhancing Cd38 transcription through IRF3 and P65. (a) WST‐8 analysis of NAD levels in 3, 10, 18, and 24 M‐BMMs ( n = 6 biological replicates). (b) Schematic of key enzymes in the NAD salvage biosynthesis pathway. (c,d) qPCR ( n = 6 biological replicates) and Western blotting ( n = 3 biological replicates) analyses of key enzymes in the NAD + salvage biosynthesis pathway in 3, 10, 18, and 24 M‐BMMs. (e) Fluorometric assay for CD38 NADase activity in 3 M‐/18 M‐BMMs ( n = 6 biological replicates). (f) WST‐8 analysis of NAD + levels in WT‐3 M‐/WT‐18 M‐BMMs and Sting ΔMφ ‐18 M‐BMMs ( n = 6 biological replicates). (g,h) Fluorometric assay and WST‐8 analysis of CD38 NADase activity (g) or NAD + levels (h) in SN‐rDPP4‐treated BMMs with or without Cgas , Sting , Irf3, and Rela knockdown ( n = 6 biological replicates). (i) Representative immunoblots of CD38 protein levels in diABZI‐treated, SN‐rDPP4‐treated, or co‐cultured BMMs with or without Cgas , Sting , Irf3, and Rela knockdown ( n = 3 biological replicates). (j) Dual luciferase reporter assay in HEK 293T cells co‐transfected with CD38 luciferase reporter, IRF3 or P65 plasmid separately, and pRL‐CMV‐Renilla ( n = 6 biological replicates). (k) Chromatin immunoprecipitation assay showing that IRF3 or P65 bound to the CD38 promoter in BMMs. Binding levels were increased in BMMs treated with SN‐rDPP4 and were abolished by STING knockdown ( n = 6 biological replicates). Data are shown as mean ± SD.

Article Snippet: Conversely, rDPP4 (Sino Biological, 10688‐H08H) administration involved once‐weekly intramedullary injections (0.2 μg per injection) for 4 weeks.

Techniques: Analysis, Western Blot, Activity Assay, Knockdown, Cell Culture, Luciferase, Transfection, Plasmid Preparation, Chromatin Immunoprecipitation, Binding Assay

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: The flow chart, differential expression analysis, ceRNA network construction, and PPI network construction. (A) The flow chart of our analysis. (B) Expression of DPP4 in 33 tumor types between normal tissues and tumor tissues. DPP4 expression was down-regulated in BRCA ( p < 0.001), CESC ( p < 0.05), CHOL ( p < 0.001), COAD ( p < 0.05), KICH ( p < 0.001), LUSC ( p < 0.001), PCPG ( p < 0.05), READ ( p < 0.01) and UCEC ( p < 0.01), while it was up-regulated in GBM ( p < 0.01), KIRC ( p < 0.001), KIRP ( p < 0.001), LIHC ( p < 0.001), LUAD ( p < 0.001), STAD ( p < 0.05) and THCA ( p < 0.001). (C) The ceRNA network of DPP4. There were three key miRNAs correlated with DPP4 and there were eight lncRNAs correlated with the key miRNAs. (D) The PPI network of DPP4. At the proteomic level, DPP4 was closely associated with FN1, CXCR4, CAV1, ITGB1, PTPRC, ADA, GCG, GIP, ACE2, and PRCP. ceRNA, competing endogenous RNA; lncRNA, long non-coding RNA; PPI, Protein-Protein Interaction. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Quantitative Proteomics, Expressing

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: Survival analysis and univariate Cox regression analysis in pan-cancer. (A) Survival analysis of DPP4 in pan-cancer. K-M survival curves indicated DPP4 was positively correlated with OS in KIRC ( p < 0.001), with DSS in KIRC ( p < 0.001), and with DFS in PRAD ( p < 0.001). (B) Univariate Cox model of OS. DPP4 expression was a risk factor in DLBC (HR = 2.757, 95%CI = 1.066-7.127, p = 0.036), LAML (HR = 2.757, 95%CI = 1.066-7.127, p = 0.036), LGG (HR = 3.474, 95%CI = 2.617-4.611, p < 0.001), and LUSC (HR = 1.118, 95%CI = 1.000-2.014, p < 0.049), and it was a protective factor in KIRC (HR = 0.787, 95%CI = 0.716-0.865, p < 0.001), KIRP (HR = 0.789, 95%CI = 0.675-0.921, p = 0.003), LUAD (HR = 0.911, 95%CI = 0.837-0.992, p = 0.031), THCA (HR = 0.701, 95%CI = 0.553-0.887, p = 0.003), and THYM (HR = 0.440, 95%CI = 0.233-0.830, p = 0.011). (C) Univariate Cox model of DFS. DPP4 expression was a risk factor in LGG (HR = 4.698, 95%CI = 1.068-20.654, p = 0.041) and UCS (HR = 1.542, 95%CI = 1.004-2.370, p = 0.048), while it was a protective factor in PRAD (HR = 0.727, 95%CI = 0.567-0.931, p = 0.012). (D) Univariate Cox model in DSS. DPP4 expression was a risk factor in LGG (HR = 3.573, 95%CI = 2.672-4.778, p < 0.001), BRCA (HR = 1.383, 95%CI = 1.133-1.689, p = 0.001), DLBC (HR = 7.111, 95%CI = 1.469-34.432, p = 0.015), and ACC (HR = 1.343, 95%CI = 1.036-1.742, p = 0.026), while it was a protective factor in KIRC (HR = 0.703, 95%CI = 0.630-0.784, p < 0.001), KIRP (HR = 0.706, 95%CI = 0.595-0.838, p < 0.001), THCA (HR = 0.593, 95%CI = 0.419-0.839, p = 0.003), and LUAD (HR = 0.872, 95%CI = 0.782-0.971, p = 0.013). (E) Univariate Cox model in PFS. DPP4 expression was a risk factor in LGG (HR = 2.442, 95%CI = 1.884-3.163, p < 0.001), PCPG (HR = 1.832, 95%CI = 1.041-3.226, p = 0.036), and LUSC (HR = 1.137, 95%CI = 1.005-1.285, p = 0.041), while it was a protective factor in KIRC (HR = 0.787, 95%CI = 0.717-0.864, p < 0.001), PRAD (HR = 0.736, 95%CI = 0.638-0.850, p < 0.001), KIRP (HR = 0.818, 95%CI = 0.711-0.942, p = 0.005), PAAD (HR = 0.794, 95%CI = 0.659-0.956, p = 0.015), and MESO (HR = 0.836, 95%CI = 0.716-0.976, p = 0.023). OS, Overall Survival; DFS, Disease-Free Survival; DSS, Disease-Specific Survival; PFS, Progression-Free Survival. *p < 0.05, **p < 0.01, ***p < 0.001 .

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: Relationship of DPP4 expression with TME in pan-cancer. (A) Correlation of DPP4 expression with MSI. It was positively correlated with MSI in COAD ( p < 0.001), ESCA ( p < 0.05), and KIRC ( p < 0.01), while it was negatively related in DLBC ( p < 0.001), HNSC ( p < 0.05), LUSC ( p < 0.001), PRAD ( p < 0.01), SKCM ( p < 0.001), and UCS ( p < 0.05). (B) Correlation of DPP4 expression with TMB. It was positively correlated with TMB in LAML ( p < 0.01), SARC ( p < 0.05), ESCA ( p < 0.001), KIRP ( p < 0.001), COAD ( p < 0.01), UCEC ( p < 0.05), GBM ( p < 0.05), LIHC ( p < 0.05), and OV ( p < 0.05), while it was negatively correlated in THYM ( p < 0.001), LUSC ( p < 0.001), CESC ( p < 0.05), PRAD ( p < 0.001), BRCA ( p < 0.001), and LUAD ( p < 0.05). (C) The relationship between DPP4 expression and immune-related genes. There is a significant association between DPP4 expression and immune-related genes across various cancers, particularly with NRP1 and HHLA2. Additionally, the majority of immune genes showed a positive correlation with DPP4 expression in BLCA, BRCA, LGG, SKCM, and THCA. (D) In PRAD, DPP4 expression was positively correlated with T cells CD4 memory resting (R = 0.18, p < 0.001), while it was negatively correlated with T cells CD8 (R = -0.19, p < 0.001), and T cells regulatory (R = -0.19, p < 0.001). MSI, Microsatellite Instability; TMB, Tumor Mutation Burden; TME, Tumor Microenvironment. *p < 0.05, **p < 0.01, ***p < 0.001 .

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Expressing, Mutagenesis

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: Drug sensitivity prediction for DPP4 in (A) CellMiner, (B) CTRP and (C) GDSC databases. The expression of DPP4 was negatively related with drug sensitivity of most drugs. However, several drugs were positively related with DPP4 expression, including perifosine and adavosertib from CellMiner, dasatinib and saracatinib from CTRP, and cetuximab and crizotinib from GDSC. (D) The molecular docking analysis of DPP4 and dasatinib. (E) The molecular docking analysis of DPP4 and midostaurin. (F) The molecular docking analysis of DPP4 and saracatinib. (G) The molecular docking analysis of DPP4 and selumetinib. The possible binding sites were illustrated. (H) RMSD values of the protein-ligand complexes over time. The DPP4-Saracatinib complex reached equilibrium after 20 ns, with its RMSD fluctuating around 2.2 Å. The DPP4-Selumetinib complex reached equilibrium after 20 ns, fluctuating around 4.1 Å. The DPP4-Dasatinib complex reached equilibrium after 20 ns, fluctuating around 2.0 Å. The DPP4-Midostaurin complex reached equilibrium after 30 ns, fluctuating around 2.2 Å. (I) Rg of the protein-ligand complexes over time. All complex systems exhibited only minor fluctuations throughout the simulation. (J) SASA of the protein-ligand complexes over time. The results indicate that the SASA of the complexes did not change significantly after ligand binding to DPP4. (K) RMSF of the protein-ligand complexes. The RMSF values for all complexes were relatively low, with most residues fluctuating below 3 Å. RMSD, Root Mean Square Deviation; RMSF, Root-Mean-Square Fluctuation; Rg, Radius of gyration; SASA, Solvent-Accessible Surface Area.

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Expressing, Binding Assay, Ligand Binding Assay, Solvent

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: A single-cell transcriptomic atlas of DPP4 in prostate cancer. (A) UMAP dimension reduction plot exhibiting 386,664 single-cell transcriptomes across nine major cell lineages (B cell, DC, endothelial cell, epithelial cell, fibroblast, mast cell, mono_macro, NK cell, and T cell) and 18 minor subtypes (luminal, basal, NE, NK cell, CD4+ T cell, CD8+ T cell, B cell, plasma cell, monocyte, macrophage, cDC1, cDC2, pDC, mast cell, fibroblast, SMC, pericyte, and endothelial cell). (B) Bubble plots depicting the feature expression of different marker genes in nine major cell subtypes. (C) UMAP dimension reduction plots by Grade1-5. (D) Bar plot demonstrating that the proportion of epithelial cells varied greatly in different grades of prostate cancer. (E) Stacked bar plots highlighting the enrichment of upregulated DEGs in epithelial cells within non-metastatic prostate cancer. (F, G) DPP4 expression was exclusively expressed in luminal cells. (H) Violin plots comparing DPP4 expression levels across ISUP grades, showing significantly higher expression in low-grade groups (p < 0.001). (I) Violin plots showing DPP4 expression across clinical T stages, indicating a significant downregulation in advanced stages (p < 0.001). (J) The interaction network illustrating the cellular communications of DPP4+ epithelial cells. (K) Heatmap summarizing the total interaction numbers, highlighting that DPP4+ epithelial cells exhibit significant communication with fibroblasts. (L) Volcano plot showing genes significantly perturbed by virtual DPP4 knockout in epithelial cells. (M) Functional enrichment analysis of the significantly perturbed genes following virtual KO of DPP4. UMAP, Uniform Manifold Approximation and Projection; AJCC, American Joint Committee on Cancer; DEG, Differential Expressed Genes; KO, Knockout.

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Single Cell, Clinical Proteomics, Expressing, Marker, Knock-Out, Functional Assay

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: Higher DPP4 expression was correlated with better prognosis in prostate cancer. (A) The inclusion and exclusion criteria of the cohort. (B) IHC scores revealed that normal tissue exhibited significantly higher DPP4 expression compared to tumor tissues ( p < 0.001). (C) Representative IHC images of both prostate cancer and normal tissues from the cohort demonstrated this difference visually. (D) The Chi square test showed that DPP4 expression was associated with WHO/ISUP grade ( p = 0.03). (E) The K-M survival curve indicated that higher DPP4 expression was not significantly correlated with OS and PFS ( p > 0.05). However, DPP4 expression tended to be a protective factor. (F) Multivariate Cox regression analysis indicated that high DPP4 expression was an independent protective factor for OS in prostate cancer patients (HR = 0.052, 95%CI = 0.0041 - 0.65, p = 0.02). IHC, Immunohistochemical; OS, Overall Survival; PFS, Progression-Free Survival.

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Expressing, Immunohistochemical staining

Journal: Frontiers in Immunology

Article Title: Integrative pan-cancer analysis of dipeptidyl peptidase 4 with clinical and in vitro validation in prostate cancer

doi: 10.3389/fimmu.2026.1616889

Figure Lengend Snippet: Dasatinib and midostaurin regulated DPP4 expression. (A) IC50 of 22Rv1 and C4–2 treated with dasatinib tested by CCK-8 assays. (B) IC50 of 22Rv1 and C4–2 treated with midostarin tested by CCK-8 assays. (C) Dasatinib treatment significantly increased DPP4 expression in C4–2 cells ( p = 0.0042, primer 1; p = 0.0029, primer 2). In contrast, midostaurin treatment reduced DPP4 expression in both cell lines (C4-2: p = 0.0218, primer 1; 22Rv1: p = 0.0172, primer 1; p = 0.0002, primer 2). IC50, Half maximal inhibitory concentration; CCK-8, Cell Counting Kit-8.

Article Snippet: Primary antibodies against human DPP4 (1:200; Affinity Biosciences Cat# DF12387, RRID: AB_2845192) were then introduced, followed by secondary antibodies conjugated with horseradish peroxidase (HRP).

Techniques: Expressing, CCK-8 Assay, Concentration Assay, Cell Counting