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

摘要

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)⁻¹i.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.