Figure 4.

CD155 is trapped in the ER of GCA Mφs

Mφs were induced from peripheral blood precursor cells as in Figure 3.

(A) Dual-color immunofluorescence imaging of Mφs stained for CD155 and cellular organelles. Markers represent the following organelles: ER, calnexin; ERGIC, P58; Golgi body, GM130; early endosome, EE1; late endosome, CD63; lysosome, LAMP2a; autophagosome, LC3; mitochondria, HSP60. Cellular stains were analyzed by imaging software to reveal co-localization as white dots (bottom panel).

(B) Co-localization coefficient between CD155 and individual cell organelles in GCA Mφs. At least 25 cells were quantified from each independent experiment.

(C) The ER fraction was isolated from healthy and GCA Mφs and CD155 protein expression analyzed by immunoblotting. Calnexin served as control. Western blot and quantitative data are shown.

(D) ER stress signatures compared in control and patient-derived Mφs. Transcript levels of ER stress markers were quantified by RT-PCR. z scores are shown (n = 12).

(E) Quantification of the ER stress-related protein BiP in control and GCA Mφs. Representative immunoblot and results from 3 independent experiments.

(F) Dual-color immunofluorescence staining of tissue sections for the Mφ marker CD163 (red) and the ER stress protein BiP (green). Nuclei marked with DAPI (blue). Temporal arteries were collected from patients with GCA. Sinonasal biopsies from patients with GPA served as disease control.

(G) Mφs were treated with the ER stress-inducer tunicamycin or DMSO. ER fractions were isolated, and CD155 was quantified by immunoblotting. Calnexin served as a control. Representative immunoblot and data from 4 independent experiments.

Mean ± SEM with individual data shown. Scale bars: 5 μm (A) and 20 μm (F). (B–E) Two-tailed unpaired t test. (G) Two-tailed paired t test. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.