Journal: Nature Communications

Article Title: CD1d-dependent rewiring of lipid metabolism in macrophages regulates innate immune responses

doi: 10.1038/s41467-022-34532-x

Figure Lengend Snippet: a qPCR expression data of Cd36 and Msr1 in WT and CD1d-KO cells ( n = 4–6). b Western-blots and quantification for CD36 and β-actin in WT and CD1d-KO BMDCs ( n = 3). c Flow cytometry images and quantification of Mean fluorescence intensity (MFI) for CD36 and MSR1 in WT and CD1d-KO cells ( n = 3–13). * p < 0.05, *** p < 0.001, one-sample t -test. d Confocal microscopy images showing staining of CD1d (red), CD36 (green) and DAPI (blue) for WT ( n = 123) and CD1d-KO BMDCs ( n = 91). Graphs show fluorescence intensity for CD1d and CD36 (left) and Pearson correlation coefficient (right). Scale bar is 20 μm. **** p < 0.0001 two-tailed unpaired t -test. e PLA was performed in WT and CD1d-KO BMDCs for CD1d and CD36. Representative images of PLA (yellow) and DAPI (blue) and quantification of PLA (puncta per cell) are shown ( n = 162–305 cells). Scale bar is 7 μm. **** p < 0.0001 two-tailed unpaired t -test. f Flow-cytometry analyses of surface CD36 in WT and CD1d-KO pMacs after incubation with oxLDL (left). MFI for CD36 (middle) or values relative to control (untreated) cells (right) are shown ( n = 6). ** p < 0.01; *** p < 0.001; **** p < 0.0001, 2-way ANOVA. g Flow-cytometry analyses of CD36 in WT pMacs pre-treated with αCD1d or isotype (iso) and incubated with oxLDL ( n = 9). CD36 levels are shown relative to control (untreated cells + isotype). * p < 0.05; paired two-tailed t -test. h Uptake of DiI-acLDL in pMacs pre-treated with αCD1d or isotype ( n = 4). ** p < 0.01, unpaired two-tailed t -test. i BMDCs were cultured with αCD1d or isotype. qPCR data show relative expression of the depicted genes ( n = 9). ** p < 0.01 one-sample t -test. j BMDCs were cultured with αCD1d or isotype (iso) and stimulated with LPS ( n = 5–9). Expression of the depicted genes was measured by qPCR. * p < 0.05, ** p < 0.01 one-sample t -test. k Uptake of DiI-acLDL in HEK293T cells transfected with CD1d, mutant CD1d (CD1d-TD or CD1d-Y) and/or CD36 ( n = 4). Some samples were incubated with αCD1d as indicated. * p < 0.05; ** p < 0.01; *** p < 0.001, one-way ANOVA. Bars in all graphs represent mean +/− SEM. Values for n represent biologically independent samples (as shown by the number of data points in each graph). n = cells isolated from individual mice ( a–c , f – j ), cells ( d , e ) or transfections ( k ). Source data are provided as a Source Data file.

Article Snippet: The plasmid containing mouse CD36 (pCMV3-HA) was purchased from SinoBiological.

Techniques: Expressing, Western Blot, Flow Cytometry, Fluorescence, Confocal Microscopy, Staining, Two Tailed Test, Incubation, Cell Culture, Transfection, Mutagenesis, Isolation

Journal: Virus research

Article Title: Select membrane proteins modulate MNV-1 infection of macrophages and dendritic cells in a cell type-specific manner

doi: 10.1016/j.virusres.2016.06.001

Figure Lengend Snippet: MNV-1 binding is reduced in CD36- and CD44-deficient bone marrow-derived dendritic cells (BMDC) but not bone marrow-derived macrophages (BMM). Wild type (WT), CD36 knockout (CD36 KO), or CD44 KO BMDC (a and c) and BMM (b and d) were infected with MNV-1 (MOI of 2). Bound virus was quantified by RT-qPCR. Results from four independent experiments performed in duplicate are shown as percent binding of virus genome equivalents to cells, which were calculated relative to the WT control cells set to 100%. *p<0.05.

Article Snippet: ELISA Binding of MNV-1 or recombinant MNV-1 Pd to the extracellular domain of recombinant murine CD36 (rCD36, catalog # 50422-M08H, SinoBiological), rCD44 (catalog # 6127-CD-050, R&D Systems), rCD98 heavy chain (rCD98, catalog # 50813-M07H, SinoBiological), and recombinant transferrin receptor 1 (rTfRc, catalog # 50741-M07H, Sino biological) was measured by ELISA.

Techniques: Binding Assay, Derivative Assay, Knock-Out, Infection, Quantitative RT-PCR

Journal: Virus research

Article Title: Select membrane proteins modulate MNV-1 infection of macrophages and dendritic cells in a cell type-specific manner

doi: 10.1016/j.virusres.2016.06.001

Figure Lengend Snippet: MNV-1 interacts with rCD36, rCD98, and rTfRc via its protruding domain (Pd). MNV-1 and MNV-1 Pd binding to the extracellular domain of recombinant proteins was evaluated by ELISA. (a) Recombinant CD36, CD44, CD98, TfRc or an irrelevant protein (BSA) were coated onto 96 well microtiter plates and incubated with MNV-1 or mock lysate. (b) Recombinant CD36, CD98, TfRc or BSA were coated onto 96 well microtiter plates and incubated with MNV-1 Pd or buffer. Results are represented as the absorbance values measured at 415 nm (A415). The dashed line is drawn at the A415 mean value for the BSA-coated wells plus twice the value of the SEM. Values above the line were considered positive. Results are from at least two independent experiments with conditions tested in duplicate.

Article Snippet: ELISA Binding of MNV-1 or recombinant MNV-1 Pd to the extracellular domain of recombinant murine CD36 (rCD36, catalog # 50422-M08H, SinoBiological), rCD44 (catalog # 6127-CD-050, R&D Systems), rCD98 heavy chain (rCD98, catalog # 50813-M07H, SinoBiological), and recombinant transferrin receptor 1 (rTfRc, catalog # 50741-M07H, Sino biological) was measured by ELISA.

Techniques: Binding Assay, Recombinant, Enzyme-linked Immunosorbent Assay, Incubation