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. 2023 Jun 17;64:102763. doi: 10.1016/j.redox.2023.102763

Fig. 2.

Fig. 2

NOX2 inhibition enhanced macrophage efferocytosis in vivo and in vitro.

A. Representative immunofluorescence staining of apoptosis and efferocytosis in carotid sections. Apoptotic cells were stained with TUNEL (green) and macrophages were stained with an anti-CD68 antibody (red). Dashed lines encircle the lumen in the first column of A. Dotted squares (the second column in A) indicate the corresponding part of the enlarged pictures (the third column in A). In the third column, yellow asterisks show examples of macrophage-associated apoptotic cells, whereas green asterisks indicate free apoptotic cells. Quantification of lesional apoptotic cells (percentage of total cells) and quantification of lesional efferocytosis index were shown in the histograms. The efferocytosis index is the free apoptotic cells/macrophage-associated apoptotic cells. n = 6 per group. *P < 0.05 versus Vehicle. B. Representative probe staining of efferocytosis in RAW264.7. Macrophage labeled by CellTrackerTM Deep Red (red) was cocultured with CFDA SE (green) labeled apoptotic Jurkat. Arrows in the fourth column of B indicates efferocytosis - macrophages with engulfed apoptotic Jurkats. C. Representative flow cytograms of efferocytosis in Vehicle and GSK2795039 group, with RAW264.7 labeled with CellTrackerTM Deep Red and Jurkats labeled with CFDA SE. Quantification of efferocytosis index in RAW264.7. The efferocytosis index is the double positive cells (Q2)/macrophage cells (Q2+ Q3) and relative to the Vehicle. n = 6 per group. *P < 0.05 versus Vehicle. D. Representative flow cytograms of efferocytosis in Vehicle and GSK2795039 group, with BMDM labeled with CellTrackerTM Deep Red and apoptotic cells labeled with CFDA SE. Quantification of efferocytosis index in BMDM. The efferocytosis index is the double positive cells (Q2)/macrophage cells (Q2+ Q3) and relative to the Vehicle. n = 6 per group. *P < 0.05 versus Vehicle. E. Representative flow cytograms of efferocytosis in Sh Control and Sh NOX2 group, with RAW264.7 labeled with CFDA SE and Jurkats labeled with CellTrackerTM Deep Red. Quantification of efferocytosis index. The efferocytosis index is the double positive cells (Q2)/macrophage cells (Q2+ Q3) and relative to the Sh Control. n = 6 per group. *P < 0.05 versus Sh Control. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)