FIG 6.

Secretory IgA is formed using pIgR after basolateral addition of dimeric Sal4 IgA. Monolayers were incubated with siRNA to knockdown pIgR expression. Results are confocal imaging of Enteroid monolayers (A–B) and HIO monolayers (C–D) stained for pIgR (red) and nuclei (blue) after incubation for 36 h with siRNA. Images shown are representative images from (n = 3) technical replicates from (n = 2) biological experimental replicates. Results show pIgR staining decreases in intensity with addition siRNA. Knockdown was further confirmed with qPCR (E). Results show relative mRNA levels of pIgR is significantly reduced with siRNA in Enteroids but not HIOs. Antibodies (10 and 20 μg/mL Sal4 dIgA) were added to monolayers basolaterally for 16 h. Apical medium was collected for a sIgA ELISA. Results show both 10 and 20 µg/mL Sal4 dIgA results in significantly more SIgA formation in control Enteroid monolayers relative to pIgR knockdowns, but not in HIOs. Confocal images shown are representative images from (n = 3) biological replicates. Bar graph depict combined data from biological replicates (n = 3) P < 0.05, one-way ANOVA with Tukey post test.