br Conclusion In conclusion our results demonstrate that

Conclusion In conclusion, our results demonstrate that incubation of both LAD and SCA rat coronary arteries rapidly increases the ETB receptor-mediated contraction as well as upregulation of ETB receptor protein levels. These findings may have important experimental implications in tissue bath experiments lasting for more than 4h, due to the possible interference of emerging ETB receptors. We demonstrate involvement of transcriptional mechanisms and ERK1/2 activity in the ETB receptor upregulation process. By using the specific MEK1/2 inhibitor U0126, the upregulation of contractile ETB receptors can be prevented by treatment with inhibitors of the central signal transduction pathways underlying the upregulation process. The here employed organ culture model represents a model situation with no-flow and serum starvation as seen in ischemic conditions. Thus, despite the obvious simplicity of the model system, our results suggest that ERK1/2-mediated upregulation of vascular contractile ETB receptors may play a pivotal role in the pathogenesis of ischemic heart diseases, and that inhibition of the underlying signal transduction may be a therapeutic strategy to avoid this pathological response of the coronary vasculature. The following are the supplementary materials related to this article.
Conflict of interest statement
Acknowledgments Thanks to Professor Karin Warfvinge (www.sciencesupport.se) and PhD Gro Klitgaard Povlsen for fruitful discussions and valuable comments on the manuscript.
Introduction Endothelin-1 (ET-1), a 21-amino AH 6809 peptide initially isolated as one of the most potent vasoconstrictors secreted by endothelial cells [52], is a member of a small family of three isopeptides (ET-1, ET-2, and ET-3) with pleiotropic activity, including cell proliferation and hormone secretion. In normal brain, the presence of ET-1 and ET-3 has been detected in endothelial cells and neurons [47]. ET-1 may also be synthesized by astrocytes during reactive gliosis, inducing astrocyte proliferation [21], and under pathological conditions such as stroke and Alzheimer's disease [53]. Hypoxia, which induces the secretion of angiogenic factors, such as vascular endothelial growth factor (VEGF), through transcriptional activation, also activates ET-1 gene transcription and results in increased ET-1 expression [46]. The biological effects of ET-1 in brain include glucose uptake [44], glutamate efflux [40], nerve growth factor expression [24], and stimulation of astrocyte proliferation [10]. Indeed, ET-1 is known to be mitogenic and anti-apoptotic for many cell types and numerous studies suggest its involvement in a variety of neoplasms [4], [32]. This hypothesis is supported by the observation of ET-1 synthesis and secretion by a variety of human cancer cells: ovarian carcinoma [3], colon cancer [1], prostate cancer [31], and melanoma [22]. Moreover, preclinical models are in favor of a contribution of ET-1 in the process of angiogenesis associated with tumor progression. Indeed, ET-1 may modulate, either directly or via VEGF production, various stages of neovascularization including proliferation, migration, and invasion of endothelial cells [32]. ET-1 pleiotropic activity is mediated by binding to specific cell surface receptors: ETA receptor (ETA-R) [2] and ETB receptor (ETB-R) [39], which belong to the G-protein-coupled, seven transmembrane domain receptor family. Pharmacological studies have shown that ETA-R preferentially binds ET-1, while ETB-R binds all three ETs with similar affinity [5]. In situ hybridization and Northern blot analyses reveal that mRNAs for rat ETA-R and ETB-R exhibit distinct cellular expression patterns both in brain and peripheral tissues. The ETA-R mRNA appears to have a restricted distribution, being predominantly expressed in vascular smooth muscle cells of peripheral tissues, bronchial smooth muscle cells, myocardium, and the pituitary gland. In contrast, the ETB-R mRNA is more widely distributed, with a prominent expression in brain, mostly in glial cells [20]. Whereas the presence of mRNA coding for components of the ET-1 system was detected in some brain tumors including meningiomas and glioblastomas [17], [33], the putative expression of ET-1 and/or its receptors in oligodendrogliomas has not yet been reported.