Notch1 regulates Sox9 expression in vitro.
(A) Relative luciferase activity in COS7 cells transfected with Col2a1 luciferase reporter along with indicated amounts of Sox9 and Notch1 intracellular domain (NICD) expression plasmids. *, p value<0.05 comparing transfection of Sox9 or NICD with empty vector; #, p value<0.05 when comparing transfection of Sox9 and NICD with either Sox9 or NICD alone. (B) Transfection of increasing amounts of NICD expression plasmid upregulates Sox9 mRNA in COS7 cells as quantified by qRT-PCR. (C) Following treatment with DAPT, Sox9 mRNA was downregulated by qRT-PCR in rat AVICs after 10 days of culture. Mean and standard deviations are shown. *, p value<0.05. (D) Immunoblot analysis showing expression of Sox9 in rat AVICs over 21 days of culture treated with DAPT or DMSO (untreated). Protein amounts were normalized using α-tubulin antibodies. Experiments in A-C were performed in triplicate and means and standard deviations are shown. Transfected DNA amounts are in nanograms.