Figure 3.

Effects of low-dose X-irradiation on adhesion of PBL to human microvascular endothelial cell (HMVEC). HMVEC were cultivated under static (A,B) or laminar (C,D) conditions for 24 h, irradiated with X-ray doses ranging from 0.1 to 0.5 Gy, and 2 Gy (C,D) or 6 Gy (A,B); stimulated with TNF-α and cocultivated with PBL 24 h afterward. The numbers of attached PBL were normalized to the averaged values obtained after treatment of HMVEC with TNF-α/0 Gy (reference value). The data analysis has been refined during the development of the assay. In initial experiments, PBL were manually counted per visual field using a microscope [(A) static and (C) laminar]. Then, a semiautomated method was developed to quantify for each field of view the numbers of both PBL and HMVEC [(B) static and (D) laminar]. N = 3 for all experiments except for (A,B): 6 Gy (N = 1); 0.2 and 0.3 Gy (N = 2); for each condition duplicates or triplicates were measured. Replicates of all experiments were pooled and mean values ± SEM were calculated. One-way ANOVA was applied; p < 0.05 was considered as significantly different from the reference value and labeled. p-values versus 0 Gy + TNF-α in detail: Panel (C): p = 0.0009 for 0.1 Gy + TNF-α; p = 0.0234 for 0.5 Gy + TNF-α and p = 0.0102 for 2 Gy + TNF-α. Panel (D): p = 0.0254 for 0.1 Gy + TNF-α; p = 0.0942 for 0.5 Gy + TNF-α and p = 0.6634 for 2 Gy + TNF-α.