Fig. 6

Tamoxifen mediated RelA deletion in skin fibroblast in vivo. a To generate fibroblast-specific RelA-/- mice, RelA CKO/CKO mice were crossed with Col1α2-CreER mice. PCR on DNA from tail biopsies was used to identify the double transgenic. Mice harboring the CKO allele were identified by the 450 bp product while those containing the WT allele were identified by the 270 bp product. b RelA^CKO/CKO Cre- and RelA ^CKO/CKO Cre + littermates were injected with 1 mg tamoxifen i.p. for 10 days. After 3 weeks, dermal fibroblasts were harvested from both genotypes and immunostained for RelA after stimulation with vehicle or TNFα (20 ng/ml). Cre + fibroblast had very little cytoplasmic RelA under basal condition and also displayed less accumulation of RelA in the nucleus after TNFα treatment. White arrows indicate cells that have decreased levels or absence of nuclear RelA. c QRT-PCR on fibroblasts isolated from Cre + (n = 4) and Cre- (n = 4) mice after tamoxifen treatment suggests significant decrease in RelA mRNA levels in Cre + fibroblasts. Data are represented as mean +/- SEM, *P < 0.04. d In addition, the same Cre + fibroblasts produce significantly less RelA protein as determined by Western blot. Data are represented as mean +/- SEM *P < 0.04. A.U is arbitrary units