Journal: Aging Cell

Article Title: Characterizing the SASP ‐Dependent Paracrine Spreading of Senescence Between Human Brain Cell Types

doi: 10.1111/acel.70673

Figure Lengend Snippet: Analysis of ligands and receptors in senescent and receiving cells. (A) Schematic depicting BulkSignalR pipeline which uses known ligand‐receptor interactions and affected downstream pathways to analyze their activation based on our bulk RNAseq data from DMSO and BrdU treated human cell lines (created with BioRender). (B) Venn diagram showing the number of receptors inferred from BulkSignalR to be activated across each of the five human cell types. Three receptors were identified in common between astrocytes (purple), endothelial cells (pink), and microglia (yellow) which were the cell types shown (Figure ) to be capable of receiving senescence signals and becoming SA β‐gal positive: CXCR7, KREMEN2, and GIPR. Only CXCR7 was expressed in the cell types capable of entering secondary senescence (astrocytes, endothelial cells, microglia) (Figure , Figure ). (C) TPM expression values of CXCR7 , its ligand CXCL12 , and DPP4 which cleaves and inactivates CXCL12 in DMSO (gray) and BrdU (red) treated cell lines ( n = 3 replicates). (D) Schematic of the four selected SASP inhibitors mechanisms of action: Bindarit is a CCL2 synthesis inhibitor which prevents p65 activation of the CCL2 gene at the promoter region, ISO‐1 is a MIF antagonist, ACT‐1004‐1239 is a CXCR7 antagonist, and Sitagliptin inhibits DPP4 preventing its action of cleaving and inactivating CXCL12 (created with BioRender). Data were analyzed by two‐way ANOVA with Tukey's multiple comparisons test (C). All graphs show mean with error bars depicting standard deviation (ns, p > 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: Treatment with DPP4 inhibitor Sitagliptin (MedChemExpress, Catalog No. HY‐13749) was used to eliminate DPP4‐dependent cleavage and inactivation of CXCL12, which is a ligand of the CXCR7 receptor.

Techniques: Activation Assay, RNA sequencing, Expressing, Standard Deviation

Journal: The Journal of Clinical Endocrinology and Metabolism

Article Title: HMGCR and Rosuvastatin Regulates GLP-1 Secretion and Expression—A Translational Study

doi: 10.1210/clinem/dgaf608

Figure Lengend Snippet: A to F, Oral glucose tolerance test (OGTT) performed after rosuvastatin (0.2 mg) treatment once daily for 27 days. A, Rosuvastatin-treated mice exhibit lower 60- and 90-minute plasma glucose levels, but B, area under the curve (AUC) was unaffected for glucose. C, Rosuvastatin-treated mice exhibit higher 5- and 60-minute plasma glucagon-like peptide 1 (GLP-1) levels, but D, AUC was unaffected for GLP-1. E, Rosuvastatin-treated mice had higher insulin secretion; AUC is presented in F. G, Long-term administration of rosuvastatin had no effect on the number of L cells in the jejunum. H to J, Long-term administration of rosuvastatin had no effect on intestinal messenger RNA expression of Gcg , Gip , Pyy , Pcsk1 , or Dpp4 in the H, duodenum; I, jejunum; or J, ileum. K, Long-term administration of rosuvastatin had no effect on β-cell mass. All parameters were assessed in 10 mice per group. Data are presented as box and whisker plots showing the minimum and maximum values (bottom and top error bars), the first (bottom of the box) and third (top of the box) quartiles and the median (middle of the box), or as a line graph showing the mean ± SEM. * P less than .05; ** P less than .01; and *** P less than .001 compared with vehicle-treated mice.

Article Snippet: RNA was reverse-transcribed using RevertAid First Strand complementary DNA synthesis kit (Thermo Scientific). qPCR for Dpp4 (Mm00494538_m1), Gip (Mm00433601_m1) Hmgcr (Mm01282499_m1), Proglucagon ( Gcg ; Mm01269055_m1), Pcsk1 (Mm00479023_m1), Pyy (Mm00520715_m1), and 2 housekeeping genes ( Hprt [Mm03024075_m1] and Tbp [Mm01277042_m1]) was performed using TaqMan Expression PCR Master Mix (Life Technologies) using the ABI Prism 7900 HT system (Applied Biosystems).

Techniques: Clinical Proteomics, RNA Expression, Whisker Assay

Journal: Advanced Science

Article Title: PET Imaging of Cardiac Inflammation in Viral Myocarditis Using a DPP4‐Targeted Probe

doi: 10.1002/advs.202516904

Figure Lengend Snippet: Pathological characterization and DPP4 expression in control and viral myocarditis (VMC) mouse hearts. (A) Typical gross images of hearts from control mice or mice with CVB3 infection (3 or 7 days post‐infection, DPI). (B) Protein levels of DPP4 in heart tissues by western blot (n = 3 per group). (C) Representative serial heart tissue sections stained with H&E, Masson's trichrome, or immunohistochemical staining for CD45. (D) Quantification of inflammatory infiltration in the heart based on H&E staining (Control and 3‐DPI groups: n = 5; 7‐DPI group: n = 10). (E) Representative immunofluorescence images showing the distribution of DPP4 protein, which largely overlapped with inflammatory lesions in the myocarditis hearts. (F) Multicolor immunofluorescence images showing DPP4 expression and co‐localization with immune cells in hearts from control and 7‐DPI VMC mice. Blue, DAPI (nucleus); Red, DPP4; Green, CD3; Yellow, CD45; Cyan, CD68. (G) Quantification of DPP4‐positive cells per high‐power field (HPF, 200x magnification) based on immunofluorescent staining (fifteen random fields per group from n = 3 mice). Each datapoint represents one biological replicate in (B and D) and one HPF in (G). Data are presented as mean ± SD. Statistical significance was assessed using one‐way ANOVA with Tukey's multiple comparisons test in (B and D) and unpaired two‐tailed Student's t‐test in (G). **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: The DPP4‐targeted probe was synthesized by conjugating the xanthine‐based DPP4 inhibitor linagliptin (MCE, USA) with the bifunctional chelator DOTA‐NHS ester (Macrocyclics, USA).

Techniques: Expressing, Control, Infection, Western Blot, Staining, Immunohistochemical staining, Immunofluorescence, Two Tailed Test

Journal: Advanced Science

Article Title: PET Imaging of Cardiac Inflammation in Viral Myocarditis Using a DPP4‐Targeted Probe

doi: 10.1002/advs.202516904

Figure Lengend Snippet: Increased DPP4 mainly originates from infiltrating immune cells. (A, B) Gene expression levels and cellular distribution of Dpp4 in hearts infected with either (A) T1L reovirus (re‐analyzed from the dataset reported by Mantri M et al.) or (B) CVB3 (re‐analyzed from Li H et al.) based on public single‐cell RNA sequencing datasets. Cell types: FB, fibroblast; EC, endothelial cell; CM, cardiomyocyte; NK, natural killer cell; pDC, plasmacytoid dendritic cell; cDC, conventional dendritic cell; Macro, macrophage; Mono, monocyte. (C–K) DPP4 expression in heart tissues from control and VMC (7 DPI) mice was analyzed by flow cytometry. (C)​Absolute number of CD45 + DPP4 + double‐positive immune cells infiltrating the hearts of VMC mice compared with control mice (n = 6 per group). (D) Representative flow cytometry histogram showing DPP4 expression on CD45 + and CD45 − cells in the heart following CVB3 infection. (E) UMAP showing different immune subsets. Bubble chart (F), bar graphs (G and H) and representative histogram (I) showing the increased DPP4 + cell proportions and the mean fluorescence intensity (MFI) in different immune subsets from control and VMC mice. Representative flow cytometry plots (J) and quantification of the frequency (K) showing co‐expression of DPP4 and the indicated activation marker (CD69, CD80 and CD86) in cardiac‐infiltrating T‐cell subsets and dendritic cells from control and VMC mice. n = 6 per group. Each datapoint represents one biological replicate in (C, G, H, and K). Data are presented as mean ± SD. Statistical significance was assessed using unpaired two‐tailed Student's t‐test in (C) and two‐way ANOVA with Tukey's multiple comparisons test in (G, H, and K). *, P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: The DPP4‐targeted probe was synthesized by conjugating the xanthine‐based DPP4 inhibitor linagliptin (MCE, USA) with the bifunctional chelator DOTA‐NHS ester (Macrocyclics, USA).

Techniques: Gene Expression, Infection, Single Cell, RNA Sequencing, Expressing, Control, Flow Cytometry, Fluorescence, Activation Assay, Marker, Two Tailed Test

Journal: Advanced Science

Article Title: PET Imaging of Cardiac Inflammation in Viral Myocarditis Using a DPP4‐Targeted Probe

doi: 10.1002/advs.202516904

Figure Lengend Snippet: Molecular docking simulations of linagliptin derivatives with DPP4. (A) Synthetic procedure of nonradioactive precursor compound DOTA‐linagliptin. (B–D) The molecular binding modes of linagliptin (B), DOTA‐linagliptin (C), and Ga‐DOTA‐linagliptin (D) with the DPP4 protein (PDB ID: 00008wku). Small‐molecule ligands are shown in blue, hydrogen bonds as colored dashed lines, and covalent bonds as solid red line.

Article Snippet: The DPP4‐targeted probe was synthesized by conjugating the xanthine‐based DPP4 inhibitor linagliptin (MCE, USA) with the bifunctional chelator DOTA‐NHS ester (Macrocyclics, USA).

Techniques: Binding Assay

Journal: Advanced Science

Article Title: PET Imaging of Cardiac Inflammation in Viral Myocarditis Using a DPP4‐Targeted Probe

doi: 10.1002/advs.202516904

Figure Lengend Snippet: Stability analysis of DPP4‐ligand complex during molecular dynamics simulations. (A) Root mean square deviation (RMSD) profiles of the DPP4‐ligand complexes. (B) RMSD profiles of ligands linagliptin, DOTA‐linagliptin, and Ga‐DOTA‐linagliptin. (C) The radius of gyration (Rg), (D) solvent‐accessible surface area (SASA), (E) root mean square fluctuation (RMSF), and (F) hydrogen bonding patterns for the three complex systems. (G‐I) Binding free energy analysis of linagliptin, DOTA‐linagliptin and Ga‐DOTA‐linagliptin with the DPP4 protein computed according to the MM/PBSA method.

Article Snippet: The DPP4‐targeted probe was synthesized by conjugating the xanthine‐based DPP4 inhibitor linagliptin (MCE, USA) with the bifunctional chelator DOTA‐NHS ester (Macrocyclics, USA).

Techniques: Solvent, Binding Assay

Journal: Advanced Science

Article Title: PET Imaging of Cardiac Inflammation in Viral Myocarditis Using a DPP4‐Targeted Probe

doi: 10.1002/advs.202516904

Figure Lengend Snippet: Targeting validation of 68 Ga‐linagliptin in vitro and in vivo. (A) Western blot and (B) immunofluorescence staining of HEK293T cells transfected with an empty vector (Control 293T) or a lentiviral vector overexpressing DPP4 (DPP4‐OE 293T). (C) Uptake of 68 Ga‐linagliptin in control and DPP4‐OE 293T cells after 60 min of incubation. (D) Representative PET images and quantification ROI result (E) of mice subcutaneously injected with control (blue arrows) or DPP4‐OE 293T cells (red arrows) in the axilla, acquired at 120 min post‐injection of 68 Ga‐linagliptin, together with the images obtained after blocking (yellow arrows). (F) In vitro dose‐response curves of linagliptin and Ga‐DOTA‐linagliptin in a DPP4 enzyme inhibition assay. IC 50 values are shown, estimated by nonlinear regression using a four‐parameter logistic (4PL) model with variable slope (Hill equation). For (C), n = 6 biological replicates per group; for (E), n = 3–5 mice per group. Each datapoint represents one biological replicate. Data are presented as mean ± SD. Statistical significance was assessed using two‐way ANOVA with Tukey's multiple comparisons test in (C) and one‐way ANOVA with Tukey's multiple comparisons test in (E). *, P<0.05, ****P<0.0001.

Article Snippet: The DPP4‐targeted probe was synthesized by conjugating the xanthine‐based DPP4 inhibitor linagliptin (MCE, USA) with the bifunctional chelator DOTA‐NHS ester (Macrocyclics, USA).

Techniques: Biomarker Discovery, In Vitro, In Vivo, Western Blot, Immunofluorescence, Staining, Transfection, Plasmid Preparation, Control, Incubation, Injection, Blocking Assay, Enzyme Inhibition Assay