Figure. 2.

Overexpression of A20 decreases inflammation in aortic allografts while increasing apoptosis in neointimal SMC. Representative IHC photomicrographs show significantly A. Decreased VCAM-1 immunostaining in the neointimal and medial layers of rAd.A20 transduced vascular allografts at 4 weeks after transplant, as opposed to saline and rAd.βgal-treated vascular allografts; This correlated with decreased infiltration of the vascular allografts by monocytes/macrophages and neutrophils, as evaluated by immunostaining with F4/80 and Gr-1 antibodies, respectively. B. Increased iNOS immunostaining in the media of rAd.A20-treated aortic allografts correlating with C increased number of TUNEL positive (blue) neointimal SMC per mm², as adjusted by Adobe scaling, when compared to saline and rAd.βgal treated allografts. Overlay of IF staining for smooth muscle cell α-actin SMA (red), Caspase 3 (green) and 4’,6-diamidino-2-phenylindole (DAPI, nuclear staining, blue) confirm that most TUNEL+ cells are neointimal SMC (yellow overlay). In A, B, and C, each bar represents mean±SE of immunostaining score, or number of F4/80, Gr-1, and TUNEL+ cells/HPF of 3-4 mice in rAd.A20-treated group and 4-7 mice in the control group. The control group represents combined saline and rAd.βgal treated vascular allografts (2-3 saline and 3-4 rAd.βgal-treated). I-intima, and M-media, original magnification X400 for VCAM-1, iNOS, TUNEL, and Caspase 3/SMA, and X200 for F4/80 and Gr-1. *p<0.05, ***p<0.001.