Figure 1
Hepatocyte-specific and conditional S100a8 and S100a9 transgene expression in TgS100a8a9hepmice. (A) Schematic representation of the transgene construct: the transcriptional transactivator (tTA) is expressed under the control of the hepatocyte-specific promoter Lap1 (TALap1), binds to and activates the tetracycline operator (TetO), which drives the simultaneous expression of S100a8-myc/his and S100a9-myc/his transgenes (TetOa8a9). Doxycycline interferes with the binding of tTA to the TetO and inhibits transgene expression. (B) Tissue-specific tgS100a8 and tgS100a9 expression was determined by semi-quantitative PCR in different tissues from TgS100a8a9hep mice using specific primers for transgenic S100a8 and S100a9 transcripts. Hprt1 levels served as control for cDNA quantity and quality. (C) TgS100a8a9hep mice were treated with (+dox) or without (-dox) doxycycline (10 μg/ml) for three weeks and transgene expression was assessed by qRT-PCR using liver cDNA. The mean value +SD of n=3 animals per group is depicted. (D) Whole liver lysates from a TgS100a8a9hep mouse (dTg) and a single TetOa8a9 transgenic (sTg) control were purified with an anti-Myc-tag IP Kit and expression of transgenic proteins was monitored by Western blot analysis using a His-tag specific antibody. HeLa cells were transiently transfected with S100a8 and S100a9 expression vectors and whole cell lysate was used as a positive control (Co). (E) S100a8/a9 protein was quantified by a heterodimer-specific ELISA in serum of Control (n=7) and TgS100a8a9hep mice (n=6); mean +SD.