Expression of VEGFR-2 during Liver Regeneration after Partial Hepatectomy in a Bioluminescence Mouse Model.

BACKGROUND: Liver regeneration requires the formation of new blood vessels. Endothelial cell proliferation is stimulated by vascular endothelial growth factor (VEGF) and its receptor tyrosine kinase VEGFR-2. The aim of this study was to investigate VEGFR-2 expression in vivo during liver regeneration after partial hepatectomy (PHx). METHODS: Transgenic VEGFR-2-luc mice were used in which the luciferase reporter gene was under control of the VEGFR-2 promoter. Following 2/3 PHx, the mice underwent in vivo bioluminescence imaging until the 14th postoperative day. Additionally, liver tissue was analyzed by immunohistochemistry, in vitro luminescence assays, and quantitative RT-PCR. RESULTS: In vivo bioluminescence imaging showed a significant increase in VEGFR-2 promoter activity after PHx. Maximum signal was recorded on the 3rd day; 8 days postoperatively the signal intensity decreased significantly. On the 14th day, bioluminescence signal reached almost baseline levels. Immunohistochemistry, quantitative RT-PCR, and in vitro luminescence confirmed a significant increase on the 3rd day following resection. The mRNA expression of VEGFR-2 was significantly higher on day 3 than preoperatively as well as on day 8. CONCLUSION: In vivo bioluminescence imaging with transgenic VEGFR-2-luc mice is feasible and provides a convenient model for noninvasively studying VEGFR-2 expression during liver regeneration. This may facilitate further experiments with modulation of angiogenesis by different substances.

In vivo imaging of antioxidant response element activity during liver regeneration after partial hepatectomy.

BACKGROUND: The nuclear factor-erythroid 2-related factor 2 (Nrf2) -antioxidant response element (ARE) pathway is important for the regulation of antioxidative stress response and detoxification. To activate the expression of its target genes, such as heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase (quinone) 1 (NQO1), Nrf2 binds to the ARE within the promoter region of these genes. Partial hepatectomy and consecutive liver regeneration lead to oxidative stress with activation of the Nrf2-ARE pathway. The aim of this study was to investigate ARE activity in vivo during liver regeneration after partial hepatectomy. MATERIALS AND METHODS: Transgenic ARE-luc mice were used. In these mice, the luciferase reporter gene is under the control of an ARE promoter element. Following 2/3 partial hepatectomy (PHx), mice underwent in vivo bioluminescence imaging up until the ninth postoperative day. In addition, liver tissue was analyzed by immunohistochemistry (Nrf2 and HO-1), quantitative reverse transcription-PCR (HO-1 and NQO1) and in vitro luminescence assays. RESULTS: Bioluminescence imaging revealed a significant increase in Nrf2-ARE activity after PHx. The signal maximum was recorded on the third day after PHx. Seven days postoperatively, the signal almost reached baseline levels. In immunohistochemistry, significantly more hepatocytes were positive for Nrf2 and HO-1 on the third postoperative day compared with baseline levels. The mRNA expression of HO-1 and NQO1 were significantly increased on day 3 as measured by qRT-PCR. CONCLUSIONS: This study demonstrated the time-dependent activation of the Nrf2-ARE system during liver regeneration in vivo. The transgenic ARE-luc mouse provided a convenient model for studying Nrf2-mediated gene expression noninvasively and may facilitate further experiments with therapeutic modulation of the antioxidative stress response.