Journal: JCI insight

Article Title: Whole-body deletion of Endospanin 1 protects from obesity-associated deleterious metabolic alterations.

doi: 10.1172/jci.insight.168418

Figure Lengend Snippet: Figure 2. Absence of Endo1 increases cell surface CD36 levels and lipid uptake in adipocytes. (A) Cell surface expression of CD36 in mature white adipo- cytes. **P < 0.01 versus WT. Results are expressed as mean ± SEM (n = 4). Two-tailed t test. (B) Total CD36 expression in gonadal adipose tissue (GAT) and

Article Snippet: Next, cells were incubated with primary Endo1 (1:500) (generated as described previously; ref. 13) and primary CD36 (1:1,000) (RD Systems, catalog AF2519), primary Endo1 (1:500) and primary 53K (1:1,000, Abcam, catalog ab27043), or primary GM130 (1:1,000, Abcam, catalog ab169276) antibodies overnight at 4°C.

Techniques: Expressing, Two Tailed Test

Journal: Journal of Alzheimer's Disease

Article Title: Cooperative Therapeutic Action of Retinoic Acid Receptor and Retinoid X Receptor Agonists in a Mouse Model of Alzheimer's Disease

doi: 10.3233/jad-132720

Figure Lengend Snippet: Fig. 3. Am80/HX630 increased NEP and IDE protein levels as well as Ide mRNA level in male APP23 mice. At 18 days after beginning treatment with Am80 (0.5 mg/kg/d) and/or HX630 (5 mg/kg/d), whole-cell extracts (A), homogenates of the membrane-bound fraction (B), and total RNA were prepared from the brains of APP23 mice (A–C). A, B) Immunoblotting with antibodies to CD36, NEP, IDE, -actin, or SRA was used to study the homogenates (40 g in A, 5 g in B). Positive controls (PC) in B are the membrane fractions (5 g) from brains of WT mice for IDE, and cell lysates from MG5 microglial cells for SRA. The upper panel in B shows representative immunoblotting results of IDE and SRA in the membrane-bound fraction. Densitometric scanning was used to quantify results. Values were normalized to SRA protein expression, and were expressed relative to the vehicle-treated mice. Error bars represent means ± SEM, n = 3 (A) and 5 (B) mice per group. ###p < 0.001 and #p < 0.05 by Dunnett’s multiple comparison test, relative to the vehicle-treated group. ∗∗p < 0.01 by Dunnett’s multiple comparison test, relative to the Am80-treated group. C) The RNA samples (1.0 g) were subjected to quantitative RT-PCR using a specific primer for Ide gene. Values were normalized to -actin gene expression. Error bars represent means ± SEM (n = 3–4). #p < 0.05 by Dunnett’s multiple comparison test, relative to the vehicle-treated APP23 mice. p = 0.062 by Student’s t-test for HX630-treated APP23 mice, relative to the vehicle-treated APP23 mice.

Article Snippet: Membranes were then incubated with antibodies to CD36 (BAF2519, 1 : 500 dilution; R&D Systems), NEP (H-321, 1 : 100 dilution; Santa Cruz Biotechnology, Santa Cruz, CA, USA), IDE (PC730, 1 : 1000 dilution; Calbiochem, Darmstadt, Germany; 9B12, 1 : 1000 dilution; Abcam, Cambridge, UK), IL-4 receptor chain (IL-4R ) (S-20, 1 : 200 dilution; Santa Cruz Biotechnology), A 1-17 (6E10, 1 : 500 dilution; Chemicon International, Temecula, CA, USA), scavenger receptor class AI/AII (SRA) (SRA-E5, 1 : 1000 dilution; a gift from Prof. M. Takeya at Kumamoto University, Kumamoto, Japan), or -actin (AC15, 1 : 5000 dilution; Sigma), followed by secondary antibodies: horseradish peroxidase-linked antibodies against rabbit, goat, or mouse Ig (each diluted 1 : 1000).

Techniques: Membrane, Western Blot, Expressing, Comparison, Quantitative RT-PCR, Gene Expression

Journal: Journal of Alzheimer's Disease

Article Title: Cooperative Therapeutic Action of Retinoic Acid Receptor and Retinoid X Receptor Agonists in a Mouse Model of Alzheimer's Disease

doi: 10.3233/jad-132720

Figure Lengend Snippet: Fig. 4. Clearance of 125I-labeled oligomeric A1-42 (o-A1-42) by rat primary microglia. A) Rat primary microglia were incubated with Am80 (5 M) and/or HX630 (5 M) for 48 h. Cell proteins (30 g) were subjected to immunoblot analysis with antibodies to CD36, IDE, or -actin. Densitometric scanning was used to quantify results. Values were normalized to -actin protein expression, and were expressed relative to those of vehicle-treated cells. Error bars represent means ± SEM (n = 3). ##p < 0.01 by Dunnett’s multiple comparison test, relative to the vehicle-treated cells. B) Cells were cultured for 48 h with Am80 (5 M) and/or HX630 (5 M) and then incubated for 6 h with 125I-labeled o-A1-42 (1 g/ml) in the presence or absence of fucoidan (100 g/ml, a scavenger receptor inhibitor). The amounts of degradation products of 125I-labeled o-A were determined. Error bars represent means ± SEM, n = 6. ###p < 0.001 and ##p < 0.01 by Dunnett’s multiple comparison test, relative to the vehicle-treated cells (column 1). ∗∗∗p < 0.001 by Student’s t-test, in the presence or absence of fucoidan. C, D) Cells were cultured for 48 h with Am80 (5 M) and/or HX630 (5 M) and then incubated for 6 h with 125I-labeled o-A1-42 (1 g/ml) in the presence or absence of fucoidan (100 g/ml), insulin (100 g/ml, an IDE inhibitor), or thiorphan (30 M, a NEP inhibitor). The amounts of cell-association (C) and degradation products (D) of 125I-labeled o-A were determined. Error bars represent means ± SEM, n = 6. ∗∗∗p < 0.001 and ∗p < 0.05 by Student’s t-test. ##p < 0.01 and #p < 0.05 for column 5 among columns 5–8, by Dunnett’s multiple comparison test.

Article Snippet: Membranes were then incubated with antibodies to CD36 (BAF2519, 1 : 500 dilution; R&D Systems), NEP (H-321, 1 : 100 dilution; Santa Cruz Biotechnology, Santa Cruz, CA, USA), IDE (PC730, 1 : 1000 dilution; Calbiochem, Darmstadt, Germany; 9B12, 1 : 1000 dilution; Abcam, Cambridge, UK), IL-4 receptor chain (IL-4R ) (S-20, 1 : 200 dilution; Santa Cruz Biotechnology), A 1-17 (6E10, 1 : 500 dilution; Chemicon International, Temecula, CA, USA), scavenger receptor class AI/AII (SRA) (SRA-E5, 1 : 1000 dilution; a gift from Prof. M. Takeya at Kumamoto University, Kumamoto, Japan), or -actin (AC15, 1 : 5000 dilution; Sigma), followed by secondary antibodies: horseradish peroxidase-linked antibodies against rabbit, goat, or mouse Ig (each diluted 1 : 1000).

Techniques: Labeling, Incubation, Western Blot, Expressing, Comparison, Cell Culture

Journal: Cell Reports

Article Title: Organotypic endothelial adhesion molecules are key for Trypanosoma brucei tropism and virulence

doi: 10.1016/j.celrep.2021.109741

Figure Lengend Snippet: Changes in expression of vascular EC receptors during a T. brucei infection (A) Expression of seven EC receptors (P-selectin, E-selectin, ICAM1, ICAM2, VCAM1, PECAM1, and CD36) measured by intravital imaging after injection of antibodies coupled to A647 fluorophore. Representative intravital images relative to g-WAT and pancreas. Scale bars: 50 μm. (B) Heatmap shows MFI values for each receptor measured in each organ on uninfected and d6-infected mice. Measurements are the mean value of at least 100 vessels measured per condition. All relevant data used to generate this figure are included in (tabs 15 and 16). See also and and .

Article Snippet: Cd36 (NM_007643) Mouse Tagged ORF Clone Lentiviral Particle , OriGENE , MR227663L4.

Techniques: Expressing, Infection, Imaging, Injection

Journal: Cell Reports

Article Title: Organotypic endothelial adhesion molecules are key for Trypanosoma brucei tropism and virulence

doi: 10.1016/j.celrep.2021.109741

Figure Lengend Snippet: Blocking specific vascular EC receptors reduced T. brucei virulence and extravasation (A) Parasitemia was followed by bioluminescence in mice treated with blocking antibodies 2 days prior to and during the initial 6 days of infection. (B) Boxplots of mean parasitemia during the first parasite peak (d5–8). (C) Survival of antibody-treated mice. ECl receptors recognized by antibodies are indicated in the key. (D) Heatmap of parasite enrichment in multiple organs on d6 post-infection, calculated from the percentage of detected parasites per area in the extravascular space, in treated versus untreated conditions. Zero was set as the baseline control, and data are plotted in a log10 scale. Each column represents a different antibody treatment of infected mice. Graphs in the middle and right panels show the respective quantitative values of parasite density in g-WAT and pancreas. All other organs are included in Figure S10 . Error bars show SD. (E) Parasitemia was followed by hemocytometer in mice homozygous (CD36 −/− ), heterozygous (CD36 +/− ), or wild-type (CD36 +/+ ) for CD36 and mice treated with blocking antibodies 2 days prior to and during the initial 6 days of infection. (F) Boxplots of the mean parasitemia during the first peak (d5–8). (G) Mouse survival as assessed in genetically modified mice. (H) Heatmap of parasite enrichment in multiple organs on day 6 post-infection, calculated from the ratio of detected parasites per area in the extravascular space, in mice homozygous or heterozygous for CD36 depletion versus wild-type littermates. Zero was set as the baseline control, and data are plotted in a log10 scale. Graphs in the middle and right panels show the extravascular parasite density in g-WAT and pancreas on day 6 post-infection. For all experiments, measurements correspond to triplicate experiments obtained from 9–12 mice. Heatmaps correspond to the score of at least 100 measurements from at least 12 individual mice. Significance relative to untreated controls is shown as ∗∗∗ p < 0.001, ∗∗ p < 0.01, or ∗ p < 0.05. Error bars show SD. All relevant data used to generate this figure are included in (tabs 17–20). See also and .

Article Snippet: Cd36 (NM_007643) Mouse Tagged ORF Clone Lentiviral Particle , OriGENE , MR227663L4.

Techniques: Blocking Assay, Infection, Control, Genetically Modified

Journal: Cell Reports

Article Title: Organotypic endothelial adhesion molecules are key for Trypanosoma brucei tropism and virulence

doi: 10.1016/j.celrep.2021.109741

Figure Lengend Snippet: CD36 facilitates T. brucei binding to and transmigration through the vascular endothelium (A) Schematic representation of static binding assay of T. brucei on plastic surfaces coated with recombinant mouse CD36 (rCD36). (B) Proportion of parasites attached to plastic surface, coated with various concentrations of recombinant mouse CD36 (rCD36). 1× PBS and 1% BSA were used as binding controls. (C) Parasite subcellular regions that bind to plastic surface. Following washouts, the proportion of parasites bound to plastic surface using the full body or the anterior end (which includes the mid-body and the free flagellum) were quantified. (D) Schematic representation of static binding assay of T. brucei on WT HUVEC and HUVECs expressing GFP or CD36. (E) Proportion of parasites attached to HUVECs (WT, GFP-, or CD36-expressing cells). Parasite attachment was quantified at six different time points (0.5, 1, 2, 4, 6, and 8 h). A statistical model considering time was performed. (F) Parasite subcellular regions that bind to HUVECs. Quantification was as described in (C). (G) Schematic representation of transwell assay of T. brucei on WT HUVEC and HUVECs expressing GFP or CD36. T. brucei were seeded on the top chamber for 1 h. (H) Proportion of parasites that migrated from the top to bottom chambers of the transwell assay (Boyden chamber). EC barrier consisted of WT HUVEC cells and HUVEC cells expressing GFP or CD36. For all panels, ANOVA tests were performed to determine significance. Significance is shown as ∗∗∗ p < 0.001, ∗∗ p < 0.01, or ∗ p < 0.05. Experiments were performed in triplicate, and 100 fields of view were quantified. Error bars show SD. All relevant data used to generate this figure are included in (tabs 21–23).

Article Snippet: Cd36 (NM_007643) Mouse Tagged ORF Clone Lentiviral Particle , OriGENE , MR227663L4.

Techniques: Binding Assay, Transmigration Assay, Recombinant, Expressing, Transwell Assay

Journal: Cell Reports

Article Title: Organotypic endothelial adhesion molecules are key for Trypanosoma brucei tropism and virulence

doi: 10.1016/j.celrep.2021.109741

Figure Lengend Snippet:

Article Snippet: Cd36 (NM_007643) Mouse Tagged ORF Clone Lentiviral Particle , OriGENE , MR227663L4.

Techniques: Purification, Blocking Assay, Control, Virus, Plasmid Preparation, Recombinant, Chemotaxis Assay, Cell Migration Assay, Software