Characterization of the renal renin-angiotensin system in transgenic mice that express rat tonin

Abstract

Introduction: Tonin is an enzyme that is able to generate angiotensin II (Ang II) from angiotensin I (Ang I) or directly from angiotensinogen. Our goal was to characterize the renal renin-angiotensin system in transgenic mice that express rat tonin (TGM`(rTon)). Materials and methods: Mice were euthanized and the kidneys removed for analysis. Tonin activity was evaluated by radioimmunoassay and angiotensin I-converting enzyme (ACE) activity by HPLC. Tonin, ACE and angiotensin II-converting enzyme (ACE2) expression was analyzed by Western blotting. Results: Tonin activity was significantly increased in TGM`(rTon) compared to their respective wild-type (WT) littermates (1.7 ± 0.21 vs 0.11 ± 0.02 nmol of Ang II/min/mg of protein). Tonin activity had a strong positive correlation with tonin expression in both TGM`(rTon) and their respective wild-type littermates. The ACE activity and expression levels of 65-kDa N-domain angiotensin I-converting enzyme isoform were significantly increased in the TGM`(rTon) when compared with WT. ACE2 expression levels were statistically significantly higher in the TGM`(rTon) when compared with WT. Angiotensin 1–7 (Ang(1–7)) and Ang I levels were significantly lower in the TGM`(rTon). Conclusions: We suggest that the environment of tonin abundance may increase N-domain ACE activity liberated by a secretase able to cleave somatic ACE.

Biochemical evaluation of the renin-angiotensin system: the good, bad, and absolute?

“…ANG II and ANG-(1-7) values derived from these methods are typically far higher in plasma (26,33,51,135), kidney (33,110,114), vasculature (48), adipose tissue (33,115), and heart (42) than by RIA, HPLC-RIA, or HPLC-MS methods (Table 1). Indeed, two recent studies quantified mouse renal ANG-(1-7) at Ͼ1,000,000 fmol/g tissue (110) and rat plasma ANG II at Ͼ300,000 pM by HPLC analysis alone (33). Rubio-Ruiz et al (115) reported an ANG-(1-7) concentration of 6,000 pM in serum and Ͼ500,000 fmol/g in adipose tissue from male Wistar rats by capillary electrophoresis.…”

Tonin Overexpression in Mice Diminishes Sympathetic Autonomic Modulation and Alters Angiotensin Type 1 Receptor Response

“…Blockade of AT1R by losartan resulted in greater BP reduction in the TGM'(rTon) group than the in the WT group; this observation indicates that AT1R signaling was preponderant in the acute control of BP in TGM'(rTon) group. Taken together, increased Ang II due to increased circulating pathway of Ang I-tonin-AngII-AT1R is one of the mechanisms of a net result of higher BP in the TGM'(rTon) mice; however, we cannot be certain that the only route for higher Ang II formation in the circulation is via tonin in the TGM'(rTon) mouse, because tonin can affect the expression of other enzymes such as ACE as we have previously shown (17).…”

“…Transgenic TGM'(rTon) mice were generated by microinjection of the rat tonin transgene into zygotes as described by Ribeiro et al (17, 18). TGM'(rTon) mice exhibited higher tonin activity in the brain, heart, kidney, liver, and bladder compared to WT (17).…”