Transcriptional regulation of the rat bradykinin B2 receptor gene: Identification of a silencer element

Kinins are involved in a variety of physiological and pathophysiological processes related to cardiovascular homeostasis, inflammation, blood flow, and nociception. Under physiological conditions, the bradykinin B2 (BKB2) receptor is constitutively expressed and mediates most of kinins' actions. However, the mechanisms regulating BKB2 receptor gene expression are still poorly understood. In this study, 4.6 kilobases of the 5′-flanking region from the rat BKB2 receptor gene were sequenced, and computer analysis revealed several sites for transcriptional factors. Nine promoter mutants were cloned in luciferase reporter gene vectors and transfected in NG108-15 cells and rat aorta vascular smooth muscle cells (VSMCs), showing several positive and negative regulatory elements. A classical silencer with 56 base pairs (bp) caused a decrease in reporter gene activity in NG108-15 cells and VSMCs and was able to inhibit the thymidine kinase promoter. Using electrophoretic mobility shift assay and surface plasmon resonance assay, protein-DNA interactions in the silencer region were determined and specific sets of protein-silencer complexes were detected in both cell types. More intense complexes were observed in the central 21 bp of the silencer and mutation in a putative SRE-1 site strongly impaired the protein-DNA binding. Down-regulation of the BKB2 receptor population in NG108-15 cells promoted by N6, 2′-O-dibutyryladenosine 3′:5′-cyclic monophosphate was paralleled by an increase in the amount of nuclear proteins bound to the silencer sequence showing an inverse relationship between protein-silencer complexes and the transcription of the BKB2 receptor gene. In summary, these data highlight the cell-specific regulation of the BKB2 receptor and the importance of a silencer element present in the regulatory region of the gene

[pt] PROGRAMA DA DISCIPLINA - ADM1480 - 2007.1

The kinin B2 receptor, which is constitutively expressed in a large number of tissues, mediates most of the known effects of bradykinin (BK). Normally undetectable in healthy tissues, the B1 receptor is strongly over-expressed under pathological conditions. BK is an important mediator in renal homeostasis and is mainly known for its natriuretic and vasodilatory effects. Recent data evidenced a role for BK in many other biological processes, such as apoptosis, development, extracellular matrix regulation and angiogenesis. In a first step to better understand how BK and its receptors could be involved in such a large variety of biological effects, we used microarray analysis to identify, under physiological conditions, the global renal gene expression profile in mice lacking either the kinin B1 or B2 receptor. Microarray experiments were performed using Agilent Mouse Oligonucleotide Microarrays (21 000 genes/microarray). Interestingly, there was a considerable number of mostly downregulated genes in both BK null mouse models compared with wild-type mice. Furthermore, a number of genes that are known to be implicated in renal physiology and/or pathology were differentially expressed in the BK null mice, which is indicative of the important role of both BK receptors in renal function

Role Of The Kinin B1 Receptor In Insulin Homeostasis And Pancreatic Islet Function

Kinins are potent vasoactive peptides generated in blood and tissues by the kallikrein serine proteases. Two distinct kinin receptors have been described, one constitutive (subtype B2) and one inducible (subtype B 1), and many physiological functions have been attributed to these receptors, including glucose homeostasis and control of vascular permeability. In this study we show that mice lacking the kinin B1 receptor (B 1 -/- mice) have lower fasting plasma glucose concentrations but exhibit higher glycemia after feeding when compared to wild-type mice. B1 -/- mice also present pancreas abnormalities, characterized by fewer pancreatic islets and lower insulin content, which leads to hypoinsulinemia and reduced insulin release after a glucose load. Nevertheless, an insulin tolerance test indicated higher sensitivity in B1 -/- mice. In line with this phenotype, pancreatic vascular permeability was shown to be reduced in B1 receptor-ablated mice. 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Delayed blockade of the kinin B1 receptor reduces renal inflammation and fibrosis in obstructive nephropathy

Renal fibrosis is the common histological feature of advanced glomerular and tubulointerstitial disease leading to end-stage renal disease (ESRD). However, specific antifibrotic therapies to slow down the evolution to ESRD are still absent. Because persistent inflammation is a key event in the development of fibrosis, we hypothesized that the proinflammatory kinin B1 receptor (B1R) could be such a new target. Here we show that, in the unilateral ureteral obstruction model of renal fibrosis, the B1R is overexpressed and that delayed treatment with an orally active nonpeptide B1R antagonist blocks macrophage infiltration, leading to a reversal of the level of renal fibrosis. In vivo bone marrow transplantation studies as well as in vitro studies on renal cells show that part of this antifibrotic mechanism of B1R blockade involves a direct effect on resident renal cells by inhibiting chemokine CCL2 and CCL7 expression. These findings suggest that blocking the B1R is a promising antifibrotic therapy