Figure 4.

β₃ Integrin is required for IκB degradation, NF-κB nuclear localization, and cIAP1 induction during PO hypertrophy. Age-matched WT and β₃^−/− mice were subjected to TAC for the indicated times (n≥4/group). A) Triton-insoluble and soluble fractions of LV were prepared and analyzed by Western blot with anti-IκB and pIκB (S32/S36) antibodies. Soluble fractions were normalized to GAPDH; insoluble fractions were normalized to actin. Quantification was conducted by densitometry of IκB level in soluble fraction by NIH Image J. Summary data show average ± se fold increase of IκB in soluble fraction of PO tissue over control. B) Fresh frozen LV tissue sections from sham and 72 h TAC mice were stained with anti-p65 NFκB antibody (green) and nuclear stain TOPRO3 (blue). Scale bar = 20 μM. C) Nuclear fractions were prepared from sham and 72 h TAC mice and were analyzed for movement of the NFκB heterodimer complex, p50:p65. Western blot analyses were performed with anti-p50 and anti-phospho-p65 (active form) NF-κB subunit antibodies. Histone H4 was used to normalize nuclear fractions from individual animals. D) Insoluble and soluble fractions were analyzed by Western blot with anti-cIAP1 antibody. Soluble fractions were normalized to GAPDH; insoluble fractions were normalized to actin.