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Timothy L Reudelhuber

Publications and source records attributed to Timothy L Reudelhuber.

18 recordsLinked to original sources
PubMed2007

Expression of an angiotensin-(1-7)-producing fusion protein in rats induced marked changes in regional vascular resistance.

We have described a transgenic rat line that expresses an angiotensin-(1-7)-producing fusion protein, the TGR(A1-7)3292. In these rats, testis acts as an angiotensin-(1-7) biological pump, increasing its plasma concentration 2.5-fold. In this study, we performed hemodynamic measurements in TGR(A1-7)3292 and age-matched Hannover Sprague-Dawley (SD) control rats, using fluorescent microspheres. Urethane-anesthetized transgenic rats had similar levels of baseline blood pressure (99 +/- 3 mmHg) as did SD rats (101 +/- 3 mmHg). However, pronounced differences were observed in other hemodynamic measurements. TGR(A1-7)3292 rats presented a significant increase in stroke volume (0.29 +/- 0.01 vs. 0.25 +/- 0.01 ml in SD), increased cardiac index (24.6 +/- 0.91 vs. 21.9 +/- 0.65 ml.min(-1).kg) and decreased total peripheral resistance (3.9 +/- 0.13 vs. 4.5 +/- 0.13 mmHg.ml(-1).min.100 g). The increase in stroke volume in transgenic rats may be partially explained by the small decrease in heart rate (326 +/- 7.0 vs. 359 +/- 6.0 beats/min in SD). Strikingly, TGR(A1-7)3292 rats presented a substantial decrease in the vascular resistance in lung, spleen, kidney, adrenals, brain, testis and brown fat tissue with no significant differences in the left ventricle, mesentery, skin, gastrocnemius muscle and white fat tissue. These results corroborate and extend previous results observed after acute angiotensin-(1-7) infusion, showing that chronic increase in circulating angiotensin-(1-7) produces sustained and important changes in regional and systemic hemodynamics. Moreover, our data suggest a physiological role for angiotensin-(1-7) in the tonic control of regional blood flow.

Adaptation, Physiological↗
PubMed2006

A hydrophobic patch in a charged alpha-helix is sufficient to target proteins to dense core secretory granules.

Many endocrine and neuroendocrine cells contain specialized secretory organelles called dense core secretory granules. These organelles are the repository of proteins and peptides that are secreted in a regulated manner when the cell receives a physiological stimulus. The targeting of proteins to these secretory granules is crucial for the generation of certain peptide hormones, including insulin and ACTH. Although previous work has demonstrated that proteins destined to a variety of cellular locations, including secretory granules, contain targeting sequences, no single consensus sequence for secretory granule-sorting signals has emerged. We have shown previously that alpha-helical domains in the C-terminal tail of the prohormone convertase PC1/3 play an important role in the ability of this region of the protein to direct secretory granule targeting (Jutras, I. Seidah, N. G., and Reudelhuber, T. L. (2000) J. Biol. Chem. 275, 40337-40343). In this study, we show that a variety of alpha-helical domains are capable of directing a heterologous secretory protein to granules. By testing a series of synthetic alpha-helices, we also demonstrate that the presence of charged (either positive or negative) amino acids spatially segregated from a hydrophobic patch in the alpha-helices of secretory proteins likely plays a critical role in the ability of these structures to direct secretory granule sorting.

Amino Acid Sequence↗
PubMed2006

Renal function in transgenic rats expressing an angiotensin-(1-7)-producing fusion protein.

Transgenic rats [TGR(A1-7)3292] present a chronic 2.5-fold increase in plasma Angiotensin-(1-7) [Ang-(1-7)] concentration. In the present study, we investigated the effects of this chronic elevation on renal function, vasopressin levels, kidney morphology, expression of Ang-(1-7) and vasopressin receptors in TGR(A1-7)3292. Urine volume and water intake were measured for 24 h. At the end of this period, plasma and urine samples were collected to evaluate renal function parameters and circulating vasopressin levels. Expression of renal V2 receptors and Mas was assessed by ribonuclease protection assay. Renal slices were processed for histological analysis. The urine flow of TGR(A1-7)3292 was significantly lower in comparison with Sprague-Dawley rats. The reduced urine volume of TGR(A1-7)3292 was accompanied by a significant increase in urinary osmolality and decrease free water clearance. Glomerular filtration rate, urinary sodium and potassium excretion were similar in both strains. No significant changes were observed in vasopressin levels as well as in V2 receptor and Mas mRNA expression in renal tissue. No changes in kidney structure of TGR(A1-7)3292 were detected. These data suggest that changes in circulating renin-angiotensin system produced by chronic increase of Ang-(1-7) levels can lead to adjustments in the water balance that are independent of vasopressin release and V2 receptor expression.

Angiotensin I↗
PubMed2005

Evidence for proprotein convertase activity in the endoplasmic reticulum/early Golgi.

Processing of precursor proteins by the proprotein convertases is thought to occur mainly in the trans-Golgi network or post-Golgi compartments. Such cleavage is inhibited by the prosegment of the convertases. During our studies of the use of the inhibitory prosegment of PC1, we noticed that a construct containing the prosegment fused to the C-terminal secretory granule sorting domain was cleaved in the endoplasmic reticulum (ER) at a pair of basic residues, best recognized by furin and PC7. This was further confirmed when this construct was fused at the C-terminus with a KDEL ER-retention signal. This suggests that the convertases could cleave some substrates within the ER, possibly by displacing the inhibitory prosegment associated with them.

Amino Acid Sequence↗
PubMed2005

Angiotensin II-dependent chronic hypertension and cardiac hypertrophy are unaffected by gp91phox-containing NADPH oxidase.

The gp91phox-containing NADPH oxidase is the major source of reactive oxygen species (ROS) in the cardiovascular system and inactivation of gp91phox has been reported to blunt hypertension and cardiac hypertrophy seen in angiotensin (Ang) II-infused animals. In the current study, we sought to determine the role of gp91phox-derived ROS on cardiovascular outcomes of chronic exposure to Ang II. The gp91phox-deficient mice were crossed with transgenic mice expressing active human renin in the liver (TTRhRen). TTRhRen mice exhibit chronic Ang II-dependent hypertension and frank cardiac hypertrophy by age 10 to 12 weeks. Four genotypes of mice were generated: control, TTRhRen trangenics (TTRhRen), gp91phox-deficient (gp91-), and TTRhRen transgenic gp91phox-deficient (TTRhRen/gp91-). Eight to 10 mice/group were studied. ROS levels were significantly reduced (P<0.05) in the heart and aorta of TTRhRen/gp91- and gp91-mice compared with control counterparts, and this was associated with reduced cardiac, aortic, and renal NADPH oxidase activity (P<0.05). Systolic blood pressure (SBP), cardiac mass, and cardiac fibrosis were increased in TTRhRen versus controls. In contrast to its action on ROS generation, gp91phox inactivation had no effect on development of hypertension or cardiac hypertrophy in TTRhRen mice, although interstitial fibrosis was reduced. Cardiac and renal expression of gp91phox homologues, Nox1 and Nox4, was not different between groups. Thus, although eliminating gp91phox-associated ROS production may be important in cardiovascular consequences in acute insult models, it does not prevent the development of hypertension and cardiac hypertrophy in a model in which the endogenous renin-angiotensin system is chronically upregulated.

Angiotensin II↗
PubMed2005

The renin-angiotensin system: peptides and enzymes beyond angiotensin II.

PURPOSE OF REVIEW: Although current pharmacotherapy for cardiovascular disease focuses on the inhibition of angiotensin II production and action, several other biologically active peptides are produced by the renin-angiotensin system. RECENT FINDINGS: Recent biochemical and physiological studies have shown that non-angiotensin II peptides, including angiotensins III, IV and 1-7, modulate blood pressure or cardiac function through novel mechanisms. Animal models support the notion that these peptides are physiologically important. SUMMARY: Although the role of these peptides has not yet resulted in the development of new therapies for the treatment of cardiovascular disease, they hold promise for identification of complimentary targets.

Aminopeptidases↗
PubMed2004

Modulation of secretory granule-targeting efficiency by cis and trans compounding of sorting signals.

Several protein domains acting through seemingly different mechanisms have been reported to have the capacity to target proteins to dense core secretory granules. Because proteins enter secretory granules with different efficiencies and because some of these proteins contain more than one granule-targeting motif, we have investigated whether compounding sorting signals could alter the efficiency of protein entry into secretory granules. In the current study we demonstrate that a paired basic cleavage site from human prorenin and an alpha-helix-containing secretory granule-sorting signal from the prohormone convertase PC1/3 can synergize to increase granule-sorting efficiency not only when located on the same protein, but also when located on distinct proteins that associate in the secretory pathway.

Animals↗
PubMed2004

Endothelium-restricted overexpression of human endothelin-1 causes vascular remodeling and endothelial dysfunction.

BACKGROUND: Endothelin (ET)-1 is a potent vasoconstrictor that contributes to vascular remodeling in hypertension and other cardiovascular diseases. Endogenous ET-1 is produced predominantly by vascular endothelial cells. To directly test the role of endothelium-derived ET-1 in cardiovascular pathophysiology, we specifically targeted expression of the human preproET-1 gene to the endothelium by using the Tie-2 promoter in C57BL/6 mice. METHODS AND RESULTS: Ten-week-old male C57BL/6 transgenic (TG) and nontransgenic (wild type; WT) littermates were studied. TG mice exhibited 3-fold higher vascular tissue ET-1 mRNA and 7-fold higher ET-1 plasma levels than did WT mice but no significant elevation in blood pressure. Despite the absence of significant blood pressure elevation, TG mice exhibited marked hypertrophic remodeling and oxidant excess-dependent endothelial dysfunction of resistance vessels, altered ET-1 and ET-3 vascular responses, and significant increases in ET(B) expression compared with WT littermates. Moreover, TG mice generated significantly higher oxidative stress, possibly through increased activity and expression of vascular NAD(P)H oxidase than did their WT counterparts. CONCLUSIONS: In this new murine model of endothelium-restricted human preproET-1 overexpression, ET-1 caused structural remodeling and endothelial dysfunction of resistance vessels, consistent with a direct nonhemodynamic effect of ET-1 on the vasculature, at least in part through the activation of vascular NAD(P)H oxidase.

Animals↗
PubMed2004

Expression of an angiotensin-(1-7)-producing fusion protein produces cardioprotective effects in rats.

Angiotensin-(1-7) [ANG-(1-7)] is a recently described heptapeptide product of the renin-angiotensin system. Because biosynthesis of ANG-(1-7) increases in animals treated with cardioprotective drugs and inactivation of the gene for angiotensin converting enzyme 2 [an enzyme involved in the biosynthesis of ANG-(1-7)] leads to the development of cardiac dysfunction, it has been suggested that ANG-(1-7) has cardioprotective properties. To directly test this possibility, we have generated transgenic rats that chronically overproduce ANG-(1-7) by using a novel fusion protein methodology. TGR(A1-7)3292 rats show testicular-specific expression of a cytomegalovirus promoter-driven transgene, resulting in a doubling of circulating ANG-(1-7) compared with nontransgenic control rats. Radiotelemetry hemodynamic measurements showed that transgenic rats presented a small but significant increase in daily and nocturnal heart rate and a slight but significant increase in daily and nocturnal cardiac contractility estimated by dP/d t measurements. Strikingly, TGR(A1-7)3292 rats were significantly more resistant than control animals to induction of cardiac hypertrophy by isoproterenol. In addition, transgenic rats showed a reduced duration of reperfusion arrhythmias and an improved postischemic function in isolated Langendorff heart preparations. These results support a cardioprotective role for circulating ANG-(1-7) and provide a novel tool for evaluating the functional role of ANG-(1-7).

Angiotensin I↗
PubMed2004

Chronic production of angiotensin IV in the brain leads to hypertension that is reversible with an angiotensin II AT1 receptor antagonist.

Angiotensin IV (Ang IV) is a metabolite of the potent vasoconstrictor angiotensin II (Ang II). Because specific binding sites for this peptide have been reported in numerous tissues including the brain, it has been suggested that a specific Ang IV receptor (AT4) might exist. Bolus injection of Ang IV in brain ventricles has been implicated in learning, memory, and localized vasodilatation. However, the functions of Ang IV in a physiological context are still unknown. In this study, we generated a transgenic (TG) mouse model that chronically releases Ang IV peptide specifically in the brain. TG mice were found to be hypertensive by the tail-cuff method as compared with control littermates. Treatment with the angiotensin-converting enzyme inhibitor captopril had no effect on blood pressure, but surprisingly treatment with the Ang II AT1 receptor antagonist candesartan normalized the blood pressure despite the fact that the levels of Ang IV in the brains of TG mice were only 4-fold elevated over the normal endogenous level of Ang peptides. Calcium mobilization assays performed on cultured CHO cells chronically transfected with the AT1 receptor confirm that low-dose Ang IV can mobilize calcium via the AT1 receptor only in the presence of Ang II, consistent with an allosteric mechanism. These results suggest that chronic elevation of Ang IV in the brain can induce hypertension that can be treated with angiotensin II AT1 receptor antagonists.

1-Sarcosine-8-Isoleucine Angiotensin II↗
PubMed2003

Expression of constitutively active guanylate cyclase in cardiomyocytes inhibits the hypertrophic effects of isoproterenol and aortic constriction on mouse hearts.

Evidence from several rodent models has suggested that a reduction of either atrial natriuretic peptide or its receptor in the heart affects cardiac remodeling by promoting the onset of cardiac hypertrophy. The atrial natriuretic peptide receptor mediates signaling at least in part via the generation of intracellular cyclic GMP. To directly test whether accumulation of intracellular cyclic GMP conveys protection against cardiac hypertrophy, we engineered transgenic mice that overexpress a catalytic fragment of constitutively active guanylate cyclase domain of the atrial natriuretic peptide receptor in a cardiomyocyte-specific manner. Expression of the transgene increased the intracellular concentration of cyclic GMP specifically within cardiomyocytes and had no detectable effect on cardiac performance under basal conditions. However, expression of the transgene attenuated the effects of the pharmacologic hypertrophic agent isoproterenol on cardiac wall thickness and prevented the onset of the fetal gene expression program normally associated with cardiac hypertrophy. Likewise, expression of the transgene inhibited the hypertrophic effects of abdominal aortic constriction, since it abolished its effects on ventricular wall thickness and greatly attenuated its effects on cardiomyocyte size. Altogether, our results suggest that cyclic GMP is a cardioprotective agent against hypertrophy that acts via a direct local effect on cardiomyocytes.

Adenylyl Cyclases↗
PubMed2002

Brain-specific restoration of angiotensin II corrects renal defects seen in angiotensinogen-deficient mice.

Mice deficient for angiotensinogen (AGT), or other components of the renin-angiotensin system, show a high rate of neonatal mortality correlated with severe renal abnormalities including hydronephrosis, hypertrophy of renal arteries, and an impaired ability to concentrate urine. Although transgenic replacement of systemic or adipose, but not renal, AGT in AGT-deficient mice has previously been reported to correct some of these renal abnormalities, the tissue target for this complementation has not been defined. In the current study, we have used a novel transgenic strategy to restore the peptide product of the renin-angiotensin system, angiotensin II, exclusively in the brain of AGT-deficient mice and demonstrate that brain-specific angiotensin II can correct the hydronephrosis and partially correct renal dysfunction seen in AGT-deficient mice. Taken together, these results suggest that the renin-angiotensin system affects renal development and function through systemically accessible targets in the brain.

Angiotensin II↗
PubMed2002

A genetically clamped renin transgene for the induction of hypertension.

Experimental analysis of the effects of individual components of complex mammalian systems is frequently impeded by compensatory adjustments that animals make to achieve homeostasis. We here introduce a genetic procedure for eliminating this type of impediment, by using as an example the development and testing of a transgene for "genetically clamping" the expression of renin, the major homeostatically responding component of the renin-angiotensin system, one of the most important regulators of blood pressure. To obtain a renin transgene whose expression is genetically clamped at a constant level, we have used single-copy chosen-site gene targeting to insert into a liver-specific locus a single copy of a modified mouse renin transgene driven by a liver-specific promoter/enhancer. The resulting transgene expresses renin ectopically at a constant high level in the liver and leads to elevated plasma levels of prorenin and active renin. The transgenic mice display high blood pressure, enhanced thirst, high urine output, proteinuria, and kidney damage. Treatment with the angiotensin II type I receptor antagonist, losartan, reduces the hypertension, albuminuria, and kidney damage, but does not affect expression of the transgene. This genetically clamped renin transgene can be used in models in which hypertension and its complications need to be investigated in a high prorenin/renin environment that is not subject to homeostatic compensations by the animal when other factors are changed.

Animals↗
PubMed2002

Tissue activity of circulating prorenin.

Both renin and its biosynthetic precursor, prorenin, are secreted into the circulation of mammals. Although the circulating levels of prorenin can exceed those of renin by as much as 100-fold in certain conditions, there is no evidence that prorenin contributes to the synthesis of circulating angiotensin peptide synthesis or increased blood pressure (BP). In the current study, we have generated a transgenic mouse line that overexpresses human prorenin in the liver and have mated these mice to a second mouse line expressing human angiotensinogen in the heart. Double-transgenic progeny display a selective increase in angiotensin I content in the heart (but not the plasma) as compared to the single-transgenic mice. These results are consistent with a model in which circulating prorenin is taken up by tissues where it can contribute to the local synthesis of angiotensin peptides. This finding may explain some of the pathologies associated with high circulating prorenin levels.

Animals↗
PubMed2002

Transendothelial transport of renin-angiotensin system components.

BACKGROUND: Vascular (interstitial) angiotensin (ANG) II production depends on circulating renin-angiotensin system (RAS) components. Mannose 6-phosphate (man-6-P) receptors and angiotensin II type 1 (AT(1)) receptors, via binding and internalization of (pro)renin and ANG II, respectively, could contribute to the transportation of these components across the endothelium. OBJECTIVE: To investigate the mechanism(s) contributing to transendothelial RAS component transport. METHODS: Human umbilical vein endothelial cells were cultured on transwell polycarbonate filters, and incubated with RAS components in the absence or presence of man-6-P, eprosartan or PD123319, to block man-6-P, AT(1) and angiotensin II type 2 (AT(2)) receptors, respectively. RESULTS: Apically applied (pro)renin and angiotensinogen slowly entered the basolateral compartment, in a similar manner as horseradish peroxidase, a molecule of comparable size that reaches the interstitium via diffusion only. Prorenin transport was unaffected by man-6-P. Apical ANG I and ANG II rapidly reached the basolateral fluid independent of AT(1) and AT(2) receptors. Basolateral ANG II during apical ANG I application was as high as apical ANG II, whereas during apical ANG II application it was lower. During basolateral ANG I application, ANG II generation occurred basolaterally only, in an angiotensin-converting enzyme (ACE)-dependent manner. CONCLUSIONS: Circulating (pro)renin, angiotensinogen, ANG I and ANG II enter the interstitium via diffusion, and interstitial ANG II generation is mediated, at least in part, by basolaterally located endothelial ACE.

Angiotensin I↗