Figure 5. Link between ctsB deficiency in mice and CD18 expression on leukocytes.

(A) CD18-associated FITC intensity on circulating mouse neutrophils in vivo measured with flow cytometry. The cells were fixed with 0.4% paraformaldehyde immediately after blood collection from the animals with cardiac puncture. n = 4 animals in each group. *P < 0.05 compared with WT mice. (B) Effect of fluid shear application (5 dyn/cm²) in the cone–plate device on CD18 expression of circulating neutrophils of WT mice measured with flow cytometry. n = 4 animals in each group. *P < 0.05 compared with unsheared group (ANOVA, Fisher’s protected least-significant difference). (C) CD18 label density ratio between sheared (5 dyn/cm²) and unsheared mouse neutrophils measured with flow cytometry. Neutrophils were treated with or without cysteine inhibitor E64 (100 μM) and ctsB-specific inhibitor CA074-me (10 µM). n = 4 animals in each group. *P < 0.05 compared with control in WT mice. (D and E) Time course of total membrane CD18 expression on neutrophils of WT mice (D) and ctsB^−/− mice (E) adhering to a glass surface as measured by laser confocal microscopy. The intensity values are normalized by the average values at 0 s and 30 s in each group. In the sheared-cell group, fluid shear (∼1.5 dyn/cm²) was applied from 60–180 s. n = 8 cells in each group. *P < 0.05 compared with values in unsheared cells at each time point.