Figure 6. The hyperosmotic upregulation of AQP5 is induced by the stimulation of gene transcription. mRNA expression was determined
with real-time RT–PCR analysis. A. Inhibition of the RNA polymerase II by actinomycin D (ActD; 5 µg/ml) abrogated the increase in AQP5 mRNA induced by hyperosmotic
(+ 100 mM NaCl) medium. The data were obtained after 6 (n=7) and 12 h (n=8) of stimulation, respectively, and are expressed
as folds of isoosmotic unstimulated control. Actinomycin D was preincubated for 30 min. B. The stability of AQP5 mRNA did not differ between isoosmotic (control; n=4) and hyperosmotic (+ 100 mM NaCl; n=5) conditions.
The cells were first treated with iso- and hyperosmotic media for 12 h, followed by the addition of actinomycin D (5 µg/ml).
Total RNA was isolated at different time periods after the addition of actinomycin D. Data are means ± SEM obtained in independent
experiments performed in triplicate, and are expressed in percentages of 0-h control. Significant difference versus isoosmotic
unstimulated control (A) and 0 h (B): *p<0.05. Significant difference versus NaCl control: ●p<0.05.