To elucidate if genetic variants in the bradykinin B2 receptor (B2) gene occur that could affect receptor expression and function, we screened for mutations in the promoter and in the coding region of the human B2 gene. In our initial study we analyzed 92 consecutive, unrelated subjects (including 25 patients with hypertrophic cardiomyopathy, 18 patients with dilated cardiomyopathy (DCM), 25 patients with hypertension, 18 patients with coronary heart disease, and 6 patients with valvular heart disease) using nonradioactive polymerase chain reaction-single-strand conformation polymorphism analysis as mutation screening method. We detected eight as yet unknown polymorphic sites in the promoter region of the B2 gene (-845 C/T, -704 C/T, -649 insG, -640 T/C, -536 C/T, -412 C/G, -143 C/T and -78 C/T) with allele frequencies between 0.5 and 13%. One of them (-412 C/G) destroys a Sp1 binding site and abolishes protein binding to this Sp1 site in human umbilical vein endothelial cells and human vascular smooth muscle cells. In the protein-coding region one new coding variant (T21M) with the potential to create a truncated receptor isoform was detected. We determined the frequency of the promoter variant at position -412 (C --> G) and the newly identified coding variant (T21M) in extended samples of 69 patients with HCM, 163 patients with DCM, 109 patients with hypertension, and 173 healthy anonymous blood donors. The promoter variant (-412 C/G) was found in one blood donor and the T21M mutation was not found in the control population. Therefore, it appears that these mutations are rare events and the determination of clinical significance will be a demanding task in the future.

Cardiovascular Diseases↗

PubMed1998

Cloning and characterization of the 5'-flanking region of the human cardiotrophin-1 gene.

To enable the analysis of the regulation of the human cardiotrophin-1 gene expression, the 5'-flanking region of the human cardiotrophin-1 gene was cloned and sequenced. Data bank search revealed several cis- active DNA elements (SP1, CREB, C/EBP, AP1 and AP-2 like and GATA) in the proximal 1.1 kb region. Six nested 5-'terminal deletion mutants from -1091/+39 to -218/+39 were fused to a luciferase reportergene and proved to be functionally active after transfection into COS-7 cells.

Animals↗

PubMed1997

Myocardial angiotensin receptor type 1 gene expression in a rat model of cardiac volume overload.

To investigate the regulation of the angiotensin receptor type 1 (AT1) in different organs in cardiac volume overload, we measured AT1 mRNA content in the atria, left and right ventricle, kidney and liver of rats with an aortocaval shunt, produced by infrarenal aortocaval puncture 4 weeks earlier. For angiotensin receptor mRNA quantitation a novel quantitative PCR procedure based on liquid phase hybridization was used that allowed the determination of absolute AT1 mRNA copy numbers and its comparison in different organs. Glyceraldehydephosphate dehydrogenase (GAPDH) mRNA was measured by RT-PCR to control externally equal mRNA content and quality of RNA extraction in shunt animals and controls. Heart weight was increased in the shunt animals, with the greatest increase in the atria. Blood pressure, plasma renin activity, plasma angiotensin I and II and aldosterone concentrations were not significantly altered. The AT1 mRNA content was significantly increased in the atria (shunt: 1167 +/- 350 copies AT1 mRNA/ng RNA vs controls: 803 +/- 240; p < 0.05). No change was found in the right or left ventricle, in the kidney and liver. The findings document that atrial hypertrophy in cardiac volume overload parallel with a significant increase in atrial AT1 mRNA content.

Animals↗

PubMed1994

Genetic polymorphisms of the angiotensin II type 1 (AT1) receptor gene.

Numerous essential, physiological effects on the cardiovascular system are attributable to angiotensin II (Ang II). Because of this we can assume that genetic changes in the specific receptor of Ang II (Ang II type 1 receptor gene, AT1) play a decisive role in the occurrence of cardiovascular disease associated with blood pressure regulation, vascular tone, cardiac and vascular growth process. To test this hypothesis, we examined the presence of polymorphisms within the coding region of the AT1 gene using polymerase chain reaction (PCR) and subsequent non-radioactive sequencing of samples from a control group with no previous history of cardiovascular complaint in individuals or immediate family. Using the Taq-sequencing procedure we found polymorphic sites, especially in the 5' region of the gene (base pair positions 9, 16, 87, 133, 186), two of which led to an exchange of the amino acid (amino acid 6: Ser<==>Pro, amino acid 45: Gly<==>Arg). Together with the silent polymorphism at base pair position 573, which our group established previously, an additional polymorphism in the 3' region of the gene was discovered. This, however, did not confer any changes in amino acid sequence. In a preliminary study we found no association between the distribution of the C/T573 polymorphic site and cardiovascular disease, such as essential hypertension (n = 20) coronary artery disease (n = 16) hypertrophic cardiomyopathy (n = 12) or dilated cardiomyopathy (n = 21). Further studies will be needed to determine to what extent the polymorphisms described are associated with cardiovascular disease.

Amino Acid Sequence↗

PubMed1994

MnlI polymorphism for the AGTR1 gene.

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Angiotensin I↗