FIGURE 1.

ZO-1 silencing impaired TER and barrier function in enterocytes. A, ZO-1 siRNA transfection decreased ZO-1 protein expression. T84 cells at 50% confluence were transfected with 200 nm of a pool of four ZO-1 or scrambled control siRNAs. Cell lysates were collected on days 4, 8, and 12 after transfection for immunoblotting for ZO-1 and β-catenin. B, ZO-1 silencing decreased TER. Cells were plated in Transwells in triplicate along with transfection reagents and 200 nm ZO-1 or control siRNAs. Medium was changed the next day and then twice a week. TER was monitored over time using a voltohmmeter. C, ZO-1 silencing impaired barrier function. Cells were grown and transfected as in B and barrier function was determined as described under “Materials and Methods” on day 16 when control siRNA-transfected cells reached high TER (>1,000 ohms·cm²). Cell lysates from each Transwell were then collected for ZO-1 immunoblotting in D. Representative immunoblots from three experiments with similar results are shown. Data from three experiments are presented as mean ± S.E., and * depicts statistical significance with a p value < 0.05 by t test.