J Physiol Volume 581, Number 3, 1173-1181, June 15, 2007 DOI: 10.1113/jphysiol.2007.131201
Cell-type specific interaction of endothelin and the nitric oxide system: pattern of prepro-ET-1 expression in kidneys of L-NAME treated prepro-ET-1 promoter-lacZ-transgenic mice
Torsten Slowinski1,2,
Philipp Kalk1,3,
Maren Christian1,
Fred Schmager4,
Katharina Relle1,3,
Michael Godes1,3,
Heiko Funke-Kaiser1,
Hans-H. Neumayer2,
Christian Bauer5,
Franz Theuring1 and
Berthold Hocher1
1 (CCR) Center for Cardiovascular Research/Institute of Pharmacology, Charité, Campus Mitte, Germany
2 Department of Nephrology, University Hospital Charité, Campus Mitte, Germany
3 Institute of Vegetative Physiology, Charite, Campus Mitte, Berlin, Germany
4 Schering AG, Berlin, Germany
5 Institute of Biochemistry and Molecular Biology, Free University, Berlin, Germany
Nitric oxide (NO) and endothelin-1 (ET-1) are known to play a major role in renal and vascular pathophysiology and exhibit a close interaction with ET-1, stimulating NO production; NO in turn inhibits ET-1 expression. Our objectives were (1) to establish a novel transgenic mouse model facilitating ET-1 expression assessment in vivo, (2) to validate this model by assessing prepro-ET-1 promoter activity in mice embryos by means of our novel model and comparing expression sites to well-established data on ET-1 in fetal development and (3) to investigate renal ET–NO interaction by assessing prepro-ET-1 promoter activity in different structures of the renal cortex in the setting of blocked NO synthases via L-NAME administration. We established transgenic mice carrying a lacZ reporter gene under control of the human prepro-ET-1 gene promoter sequence (8 kb of 5' sequences). Bluo-Gal staining of tissue sections revealed intracellular blue particles as indicators of prepro-ET-1 promoter activity. In mouse embryos, we detected high prepro-ET-1 promoter activity in the craniofacial region, as well as in bone and cartilage consistent with the literature. In order to investigate the interaction of ET-1 and NO in the kidney in vivo, transgenic mice at the age of 3–4 months were treated with a single dose of the NO synthase inhibitor L-NAME (25 mg (kg bw)–1
I.P.) 12 h before kidney removal. Bluo-Gal staining of kidney sections revealed intracellular blue particles as indicators of prepro-ET-1 promoter activity in tubular and vascular endothelium and glomerular cells. Particle count was closely correlated to kidney tissue ET-1 content (R
= 0.918, P < 0.001). Comparison of counts revealed an increase by 135 ± 53% in L-NAME treated (n
= 12) compared to non-treated mice (n
= 10, P
= 0.001). Cell-type specific evaluation revealed an increase of 136 ± 51% in tubular (P
= 0.001) and 105 ± 41% in glomerular cells (P
= 0.046), but no significant increase in vascular endothelium. In conclusion, our study revealed a close interaction of renal endothelin and the NO system in a cell-type specific manner. Our new transgenic model provides a unique opportunity to analyse regulation of the ET system on a cellular level in vivo.
(Received 27 February 2007;
accepted after revision 23 March 2007;
first published online 29 March 2007)
Corresponding author B. Hocher: Center for Cardiovascular Research/Institute of Pharmacology, Charité Mitte, Hessische Str. 3-4, 10115 Berlin, Germany. Email: berthold.hocher{at}charite.de; homepage: http://www.ccr.charite.de/site/html/de/ag_hocher.html
T. Slowinski and P. Kalk contributed equally as first authors. F. Theuring and B. Hocher contributed euqually as senior authors to this work.
Copyright © 2007 The Physiological Society.
