Detection of CD28/CD86 co‐stimulatory molecules and surface properties of T and dendritic cells: An AFM study

Abstract

Although the importance of B7/CD28 co-stimulation has been widely studied, little is known about their nano-spatial localization and their corresponding cells' biophysical and biomechanical properties. Here, we investigated the morphological, biophysical, and biomechanical properties of T cells and dendritic cells (DCs) by atomic force microscopy (AFM) and force curves. The nano-spatial distribution of CD28 and CD86 antigen on T cells and DCs was detected by CD86 or CD28 antibody-functionalized AFM tip. Single-molecule force spectroscopy (SMFS)-based force volumes and quantum dots (QDs)-based fluorescence imaging demonstrated that the co-stimulatory molecules were not randomly distributed over the cells' surface, but more than 80% of CD28 and CD86 molecules appeared to be expressed as 100-200 nm nanoclusters and polarize dominantly in the peak of the cell membrane fluctuations. AFM imaging and quantitative analysis showed that the roughness of mature DCs (mDCs) was higher than that of immature DCs (iDCs). The adhesion force distribution of iDCs and mDCs was heterogeneous while the elasticity distribution was homogeneous locally. In addition, mDCs had a fourfold increase of Young's modulus of iDCs, indicating the contribution of the actin cytoskeleton to the elastic properties of the cells. Taken together, the nano-cluster distribution of CD28 and CD86, the rough mDCs surface, the higher adhesion force and elasticity of mDCs may facilitate to the occurrence of B7/CD28 co-stimulation signals and the formation of immune synapse. These nanoscale findings provide new insights into the antigen-presenting function of DCs, the T cell activation and ultimate immune response. SCANNING 38:365-375, 2016. © 2015 Wiley Periodicals, Inc.

Periodic Arrangement of Lipopolysaccharides Nanostructures Accelerates and Enhances the Maturation Processes of Dendritic Cells

“…As it stretched out, the height and width gradually decreased to 17 nm. The height and width of individual dendrites were consistent with previous AFM study of mature BMDCs using sLPS presentation, while more and longer dendrites were present for BMDCs activated by nanostructures of LPS.…”

“…First, we compared the measurements from LSCM and flow cytometry using sLPS activation of BMDCs. In LSCM measurements, three dyes were selected for ICC staining (see Materials and Methods for details): , Hoechst 33342 to highlight nucleus; anti-CD11c-615 to recognize CD11c, a preferential integrin in DC; and anti-CD86-488 to highlight CD86 expression. The first two dyes marked viable BMDCs, while anti-CD86-488 revealed the cells with detectable expression of CD86.…”

Effects of 2,3,7,8-Tetrachlorodibenzo-p-dioxin on T Cell Differentiation in Primary Biliary Cholangitis

“…As shown in Figure 2, the MFI of MHC II (a), CD86 (b), CD80 (c), and CD40 (d) increased in the TCDD group compared with the control and TH+TCDD groups in both patients with PBC and HCs (p < 0:05). In addition, CH223191 as an antagonist of dioxin-induced AhR activation suggested that DCs were activated by AhR [16].…”