FIGURE 5.

Short-time treatment with PD-L1 decreases KCa3.1 activity in a calmodulin-independent manner. (A) Flow cytometry histogram and geometric mean fluorescence intensity (gMFI) values (B) for CaM expression in activated CD8⁺ PBTs from HD donors (n = 3) in the absence and presence of PD-L1. (C) Representative recordings of KCa3.1 currents in activated CD8⁺ PBTs from HDs showing the effect of PD-L1 (PD-L1-Fc, 10 μg/ml) and CaM (50 µM). (D) Average normalized KCa3.1 conductance (G, nS) measured in the absence and presence of PD-L1, with and without CaM. All conductance values are normalized to average conductance value obtained from control recordings. Cells were pre-incubated with plate-bound PD-L1 (PD-L1-Fc,10 μg/ml, for 72 h) activated using anti-CD3/CD28 antibodies and treated with or without CaM (50 µM), that was delivered intracellularly via patch pipette during recordings (n = 15–18 cells per group from 3 HDs). The values in panel (B) are represented as bar graphs. Each symbol represent an individual HD. The values are represented as mean ± SEM. The values in panel (D) are represented as box and whisker plots. The lower and upper bound of the box represent 25^th and 75^th percentiles respectively. Median values are shown as horizontal line. The lower and upper error bars represents 10^th and 90^th percentile respectively. Data in panel (B) were analyzed by t-test and data in panel (D) were analyzed by ANOVA on ranks (p = 0.008) with Dunn’s post hoc analysis.