Urinary Angiotensinogen Excretion Research Articles

Effect of losartan on proteinuria and urinary angiotensinogen excretion in non-diabetic patients with chronic kidney disease

PurposeActivation of the rennin–angiotensin system (RAS) is thought to contribute to hypertension and proteinuria, and eventually to the progression of chronic kidney disease (CKD). Recent evidence suggests that urinary angiotensinogen...

Salt-induced renal injury in SHRs is mediated by AT1 receptor activation

This study aimed to examine the effects of salt loading, with or without simultaneous angiotensin receptor blocker (ARB) treatment, on the systemic and tissue renin-angiotensin system (RAS) in spontaneously hypertensive rats (SHRs). Evaluation was performed early (4 weeks) in the course of salt loading in order to examine initial mediating events of cardiovascular and renal damage produced by salt excess. Four groups of rats were studied. Group 1 received regular rat chow (normal-salt diet); group 2 received normal-salt diet and an ARB (losartan, 30 mg/kg per day); group 3 received high-salt (8%) chow; and group 4 received high-salt diet and losartan. High-salt diet increased systolic pressure to 193±1 mmHg compared to 180±2 in normal-salt diet group. Losartan reduced SBP in SHRs fed normal-salt diet but did not reduce SBP in the SHRs fed high-salt diet (192±2 mmHg). High-salt diet markedly increased urinary protein excretion from 27±4 to 64±13 mg/day and this increase was ameliorated by losartan (40±9 mg/day). In SHRs on high-salt diet, plasma angiotensin II concentration increased three to four-fold, whereas urinary angiotensinogen excretion increased 10-fold; and these changes were significantly reduced by losartan. High-salt diet accelerated glomerular injury and interstitial fibrosis in SHRs which were reduced by losartan. These results demonstrate that the activity of RAS was either not suppressed or, even augmented, after 4 weeks of salt loading despite high salt intake and increased SBP. The data suggest that an augmented intrarenal RAS during high-salt diet may contribute to the development of renal injury in this experimental model.

Comments on Point:Counterpoint: The dominant contributor to systemic hypertension: Chronic activation of the sympathetic nervous system vs. Activation of the intrarenal renin-angiotensin system

Comments on Point:Counterpoint: The dominant contributor to systemic hypertension: Chronic activation of the sympathetic nervous system vs. Activation of the intrarenal renin-angiotensin system

Enhancement of renin and prorenin receptor in collecting duct of Cyp1a1-Ren2 rats may contribute to development and progression of malignant hypertension

To determine whether in the transgenic rat model [TGR(Cyp1a1Ren2)] with inducible ANG II-dependent malignant hypertension changes in the activation of intrarenal renin-angiotensin system may contribute to the pathogenesis of hypertension, we examined the gene expression of angiotensinogen (AGT) in renal cortical tissues and renin and prorenin receptor [(P)RR] in the collecting duct (CD) of the kidneys from Cyp1a1Ren2 rats (n = 6) fed a normal diet containing 0.3% indole-3-carbinol (I3C) for 10 days and noninduced rats maintained on a normal diet (0.6% NaCl diet; n = 6). Rats induced with I3C developed malignant hypertension and exhibited alterations in the expression of renin and (P)RR expressed by the CD cells. In the renal medullary tissues of the Cyp1a1Ren2 transgenic rats with malignant hypertension, renin protein levels in CD cells were associated with maintained renin content and lack of suppression of the endogenous Ren1c gene expression. Furthermore, these tissues exhibited increased levels of (P)RR transcript, as well as of the protein levels of the soluble form of this receptor, the s(P)RR. Intriguingly, although previous findings demonstrated that urinary AGT excretion is augmented in Cyp1a1Ren2 transgenic rats with malignant hypertension, in the present study we did not find changes in the gene expression of AGT in renal cortical tissues of these rats. The data suggest that upregulation of renin and the s(P)RR in the CD, especially in the renal medullary tissues of Cyp1a1Ren2 transgenic rats with malignant hypertension, along with the previously demonstrated increased availability of AGT in the urine of these rats, may constitute a leading mechanism to explain elevated formation of kidney ANG II levels in this model of ANG II-dependent hypertension.

Enhanced Urinary Angiotensinogen Excretion in Cyp1a1-Ren2 Transgenic Rats With Inducible ANG II-Dependent Malignant Hypertension

IntroductionPrevious studies have demonstrated that the urinary excretion of angiotensinogen is significantly increased in ANG II-infused hypertensive rats, which is associated with an augmentation of intrarenal ANG II levels. These findings suggest that urinary angiotensinogen excretion rates provide an index of intrarenal ANG II levels in ANG II-dependent hypertensive states. However, little information is available regarding the urinary excretion of angiotensinogen in ANG II-dependent malignant hypertension. MethodsThis study was performed to determine if urinary angiotensinogen excretion is increased in Cyp1a1-Ren2 transgenic rats [strain name: TGR(Cyp1aRen2)] with inducible ANG II-dependent malignant hypertension. Adult male Cyp1a1-Ren2 rats (n=6) were fed a normal diet containing 0.3% indole-3-carbinol (I3C) for 10days to induce ANG II-dependent malignant hypertension. ResultsRats induced with I3C exhibited pronounced increases in systolic blood pressure (208 ± 7 versus 127 ± 3mm Hg; P < 0.001), marked proteinuria (29.4 ± 3.6 versus 5.9 ± 0.3mg/d; P < 0.001) and augmented urinary angiotensinogen excretion (996 ± 186 versus 241 ± 31 ng/d; P < 0.01). Chronic administration of the AT1 receptor antagonist, candesartan (25mg/L in drinking water, n=6), prevented the I3C-induced increases in systolic blood pressure (125 ± 5mm Hg; P < 0.001), proteinuria (7.3 ± 1.0mg/d; P < 0.001) and urinary angiotensinogen excretion (488 ± 51 ng/d, P < 0.01). ConclusionsThese data demonstrate that the urinary excretion of angiotensinogen is markedly augmented in ANG II-dependent malignant hypertension. Such increased urinary angiotensinogen excretion may contribute to augmented intrarenal ANG II levels and, thereby, to the increased blood pressure in Cyp1a1-Ren2 transgenic rats with inducible ANG II-dependent malignant hypertension.

Enhanced Urinary Angiotensinogen Excretion in Cyp1a1‐Ren2 Transgenic Rats with Inducible ANG II‐Dependent Malignant Hypertension

The present study was performed to determine if urinary angiotensinogen excretion is increased in Cyp1a1‐Ren2 transgenic rats [strain name: TGR(Cyp1aRen2)] with inducible ANG II‐dependent malignant hypertension. Adult male Cyp1a1‐Ren2 rats (n=6) were fed a normal diet containing 0.3% indole‐3‐carbinol (I3C) for 10 days to induce ANG II‐dependent malignant hypertension. Rats induced with I3C exhibited pronounced increases in systolic blood pressure (SBP) (208±7 vs. 127±3 mmHg, P&lt;0.001), marked proteinuria (29.4±3.6 vs. 5.0±0.4 mg/day, P&lt;0.01), and augmented urinary angiotensinogen excretion (996±186 vs. 199±19 mg/day, P&lt;0.05). Chronic administration of the AT1 receptor antagonist, candesartan (25 mg/L in drinking water, n=6), prevented the I3C‐induced increases in SBP (125±5, P&lt;0.001), proteinuria (7.3±1.5 mg/day, p&lt;0.001) and urinary angiotensinogen excretion (488±51 mg/day, P&lt;0.05). The present findings confirm that AT1 receptor activation contributes to the increase in SBP and proteinuria in Cyp1a1‐Ren2 transgenic rats with ANG II‐dependent malignant hypertension. These data also demonstrate that the urinary excretion of angiotensinogen is markedly augmented in ANG II‐dependent malignant hypertension. Such increased urinary angiotensinogen excretion likely reflects an AT1 receptor‐mediated stimulation of intrarenal angiotensinogen production which may contribute to augmented intrarenal ANG II levels and, thereby, to the increased blood pressure in Cyp1a1‐Ren2 transgenic rats with inducible ANG II‐dependent malignant hypertension.

Urinary excretion of angiotensinogen in male and female Sprague Dawley rats during angiotensin II infusion and high salt diet

We have found that urinary angiotensinogen (AGT) excretion (uAGT) differs between male(♂)and female(♀) Sprague Dawley (SD) rats on a normal salt diet (♂: 60±26 vs ♀: 4.5 ±0.2ng/day, p&lt;0.05). In this study, we examined if the effects of a high salt (HS) diet on uAGT exhibits sexual dimorphism during Angiotensin (ANG) II dependent hypertension. Four groups of age‐matched (8 weeks) male (n= 20) and female (n=17) SD rats were studied: 1) high salt diet (HS; 8%NaCl); 2) ANG II (80ng/min, sc; for 14 days); 3) ANG II + HS; 4) ANG II + HS + Candesartan. (CAND, 25 mg/L in drinking water). uAGT levels were measured by enzyme immunoassay. HS diet alone did not raise uAGT levels (♂: 107±23 vs. ♀: 5±2ng/day) from baseline; however, AGT increased significantly during ANG II infusion (♂: 1107±1061 vs ♀: 206±74ng/day, p&lt;0.01) and was augmented even further in ANG II + HS (♂: 4363±1673 vs. ♀: 8022±1300ng/day, p=ns). Candesartan ameliorated this effect with a better response in female than in male rats.(♂: 240±28 vs. ♀: 10±4 ng/day, p&lt; 0.01). The results demonstrate a sexual dimorphism in the uAGT excretion rate during baseline conditions and HS diet characterized by higher levels in the males. The augmentation of uAGT with HS + ANG II is ameliorated more effectively by ARB treatment in female ANG II hypertensive rats than in males.NIH (P20 RR 017659), Tulane‐BIRCWH (K12HD043451)

High salt intake exacerbates renal tissue oxidative stress and urinary angiotensinogen excretion during AngII‐dependent hypertension.

High salt (HS) intake inhibits systemic and intrarenal RAS; however, when AngII is not suppressed HS exacerbates hypertension and renal injury. We investigated the effects of HS on the magnitude of intrarenal oxidative stress associated with angiotensinogen urinary excretion (uAGT) in chronic AngII‐infused rats. In Sprague‐Dawley rats: 1) Control, normal salt diet [NS, n=5]; 2) HS diet [8% NaCl, 12 d, n=5]; 3) AngII‐infused in NS rats [AngII, 80 ng/min, 14 d, n=5] and 4) AngII‐infused in HS rats [AngII+HS, n=5] we measured systolic blood pressure (SBP), proteinuria, NADPH oxidase activity, n‐Tyr residues and uAGT. HS alone did not change SBP, but led to hypertension when combined with AngII. In contrast, HS alone led to proteinuria that was further increased in HS+AngII rats. The uAGT excretion rates increased in AngII+HS rats compared to AngII rats (AngII=107±18 vs AngII+HS=4363±1296 ng/day). Renal tissue oxidative stress measured by n‐Tyr Western blot increased in both HS and AngII+HS rats; however the NADPH oxidase activity increased only in HS+AngII rats (HS=10,674±2819; AngII+HS=18,663±1328 RLU/mg protein). HS exacerbates renal tissue oxidative stress during AngII‐dependent hypertension which might be associated with increased uAGT, proteinuria, and blood pressure in AngII‐infused rats.NIH [IdeA Program (P20RR01765906); Tulane‐BIRCWH Program (K12HGO4351)] and AHA (09PRE2209079; 09BGIA2280440)

Increased Urinary Renin Excretion Rate in Chronic Ang II‐infused Rats Fed a High Salt Diet leads to augmented urinary Ang II levels.

This study examined the urinary excretion of renin (uRen), angiotensinogen (uAGT), and angiotensin II (uAngII) in chronic AngII‐infused Sprague‐Dawley rats fed a high salt (HS) diet. Three groups were studied: 1) AngII‐infused [80 ng/min, 14 d, n=5]; 2) AngII‐infused plus HS [Ang II+HS; 8% NaCl; n=4]; 5) AngII‐infused, HS plus an AngII type 1 receptor blocker (ARB) [Ang II+HS+Candesartan, 25 mg/L, drinking water, 14 d, n=5]. Systolic blood pressures increased in AngII (175±10 mmHg) and were augmented by HS in AngII+HS rats (222±8 mmHg). By day 13, uRen, uAGT and uAngII excretion rates increased in AngII‐infused rats, and in AngII+HS rats: [uRen (AngII: 9±3 vs. AngII+HS: 10±4 ×10‐5 Enzyme Units/day); uAGT (AngII: 1107±106 vs. AngII+HS: 4363±1673 ng/day); uAngII: (HS:1,851±327; AngII: 3,813±431; AngII+HS: 8,570±889 vs. NS: 2080±361 fmol/day; respectively)]. ARB treatment prevented these increases (uRen: 5±3×10‐5 Enzyme Units/day; uAGT: 240±28 ng/day; uAngII: 3,796±1,059 fmol/day). Thus, HS exacerbates the increases in uRen, uAGT, and uAngII as well as the systolic blood pressure in AngII‐dependent hypertensive rats.NIH [IdeA Program (P20‐RR‐017659‐06) and the Tulane‐BIRCWH Program (K12HD043451)]; and the AHA (09PRE2209079 and 09BGIA2280440).

Urinary Renin Excretion is augmented in Chronic Angiotensin II‐infused Sprague‐Dawley Hypertensive Rats

Renin is upregulated in principal cells of the collecting ducts of angiotensin II (AngII)‐dependent hypertensive rats; however it remains unclear if increased distal nephron‐derived renin is secreted into the urine during AngII‐dependent hypertension. Urinary renin (uRen), angiotensinogen (uAGT) and AngII (uAngII) excretion was measured in chronic AngII‐infused male Sprague‐Dawley rats [80 ng/min, SC minipumps for 14 d, n=5] and control, sham‐operated rats [n=5]. Systolic blood pressures increased in the AngII rats by Day 5 and continued to increase throughout the study (Day 13; AngII: 175±10 vs. sham: 116±2 mm Hg). Although plasma renin activity was suppressed in the AngII‐infused rats (AngII: 0.3±0 vs. sham: 5.5±2 ng Ang I/ml/h); renin content in renal medullary tissues did not change (AngII: 10,564±1,476 vs. sham: 9,438±1,588 ng Ang I/h/mg; p=ns). Excretion of uAGT and uAngII increased in the AngII rats compared to sham rats, respectively [uAGT (AngII:1107±106 vs. sham:60±26 ng/day); uAngII (AngII: 3813±431 vs. sham: 2080±361 fmol/day). Importantly, uRen excretion rate at Day 13 was markedly increased in the AngII rats (AngII: 9±3 vs. sham: 1±1×10−5 Enzyme Units/day; p&lt;0.05). Augmented uRen and uAGT may contribute to increased intrarenal AngII formation in AngII‐dependent hypertensive rats.NIH [IdeA (P20‐RR‐017659‐06), Tulane‐BIRCWH (K12HD043451)]; and AHA (09PRE2209079 and 09BGIA2280440).

Enhanced Distal Nephron Sodium Reabsorption in Chronic Angiotensin II–Infused Mice

Chronic angiotensin II (Ang II) infusions enhance urinary excretion of angiotensinogen, suggesting augmentation of distal nephron sodium reabsorption. To assess whether chronic Ang II infusions (15 ng/min for 2 weeks) enhance distal nephron sodium reabsorption, we compared sodium excretion before and after blockade of the 2 main distal nephron sodium transporters by IV amiloride (5 mg/kg of body weight) plus bendroflumethiazide (12 mg/kg of body weight) in male C57/BL6 anesthetized control mice (n=10) and in chronic Ang II-infused mice (n=8). Chronic Ang II infusions increased systolic blood pressure to 141+/-6 mm Hg compared with 106+/-4 mm Hg in control mice. After anesthesia, mean arterial pressure averaged 97+/-4 mm Hg in chronic Ang II-infused mice compared with 94+/-3 mm Hg in control mice, allowing comparison of renal function at similar arterial pressures. Ang II-infused mice had lower urinary sodium excretion (0.16+/-0.04 versus 0.30+/-0.05 microEq/min; P<0.05), higher distal sodium reabsorption (1.74+/-0.18 versus 1.12+/-0.18 microEq/min; P<0.05), and higher fractional reabsorption of distal sodium delivery (91.1+/-1.8% versus 77.9+/-4.3%; P<0.05) than control mice. Urinary Ang II concentrations, measured during distal blockade, were greater in Ang II-infused mice (1235.0+/-277.2 versus 468.9+/-146.9 fmol/mL; P<0.05). In chronic Ang II-infused mice treated with spironolactone (n=5), fractional reabsorption of distal sodium delivery was similarly augmented as in chronic Ang II-infused mice (94.6+/-1.7%; P<0.01). These data provide in vivo evidence that there is enhanced distal sodium reabsorption dependent on sodium channel and Na(+)-Cl(-) cotransporter activity and increased urinary Ang II concentrations in mice infused chronically with Ang II.

ACTIVATION OF REACTIVE OXYGEN SPECIES AND THE RENIN–ANGIOTENSIN SYSTEM IN IgA NEPHROPATHY MODEL MICE

1. Although IgA nephropathy is the most common form of primary glomerulopathy, the detailed mechanisms underlying its development remain uncertain. 2. In the present study, we used male high IgA strain of ddY (HIGA) mice as the IgA nephropathy model and age-matched male BALB/c mice as the control. Recent studies have demonstrated that reactive oxygen species (ROS)-dependent enhancement of the renin-angiotensin system (RAS) plays a potential role in the development and progression of renal injury. Therefore, in the present study we periodically measured the systolic blood pressure (SBP) of mice over the period 21-25 weeks of age and estimated markers for ROS, RAS and renal damage after mice had been killed at 25 weeks of age. 3. Markers for ROS (urinary 8-isoprostane excretion and renal 4-hydroxy-2-nonenal accumulation), RAS (renal angiotensinogen protein expression, urinary angiotensinogen excretion and renal angiotensin II) and renal damage (desmin-positive area and urinary protein excretion), as well as SBP, were significantly increased in HIGA mice compared with control BALB/c mice. 4. The data suggest that both ROS and the RAS are activated at an early phase in IgA nephropathy model mice.

Angiotensin II Type 1 Receptor Blockers Reduce Urinary Angiotensinogen Excretion and the Levels of Urinary Markers of Oxidative Stress and Inflammation in Patients with Type 2 Diabetic Nephropathy

To demonstrate that the administration of an angiotensin (Ang) II type 1 receptor (AT1R) blocker (ARB) inhibits the vicious cycle of high glucose (HG)-reactive oxygen species (ROS)-angiotensinogen (AGT)-Ang II-AT1R-ROS by suppressing ROSs and inflammation, thus ameliorating diabetic nephropathy (DN). Thirteen hypertensive DN patients were administered ARBs, and the following parameters were evaluated before and 16 weeks after the treatment: urinary AGT (UAGT), albumin (albumin-creatinine ratio: ACR), 8-hydroxyde-oxyguanosine (8-OHdG), 8-epi-prostaglandin F2alpha(8-epi-PGF2alpha), monocyte chemoattractant protein (MCP)-1, interleukin (IL)-6, and IL-10. ARB treatment reduced the blood pressure and urinary levels of AGT, ACR, 8-OHdG, 8-epi-PGF2alpha, MCP-1, and IL-6 but increased the urinary levels of IL-10. The reduction rate of UAGT correlated with the reduction rate of blood pressure; the reduction rates of the urinary ACR, 8-OHdG, 8-epi-PGF2alpha, MCP-1, and IL-6 levels; and the increase rate of the urinary IL-10 levels. Moreover, subjects who had high UAGT values at baseline exhibited higher reduction rates of urinary albumin excretion. ARB-induced blockade of the abovementioned vicious cycle contributes to the renoprotective effects of ARBs in DN. The urinary levels of AGT could represent a predictive factor for reduced ACR in patients receiving ARB treatment.

Intrarenal RAS activity and urinary angiotensinogen excretion in anti-thymocyte serum nephritis rats

The differential roles of circulating and intrarenal renin-angiotensin system (RAS) in glomerulonephritis have not been elucidated. In this study, we investigated the levels of circulating and intrarenal RAS activity and urinary angiotensinogen (AGT) excretion in anti-thymocyte serum (ATS) nephritis induced by an ATS injection (ATS group). The effect of olmesartan, an angiotensin II (ANG II) type 1 receptor blocker (ARB), on the development of nephritis was also examined (ATS+ARB group). In addition, the rats received a saline injection instead of ATS (control group). Mesangial proliferation with transient proteinuria, which peaked at day 7, was significantly increased in the ATS group compared with the control group. The levels of glomerular AGT mRNA, intrarenal ANG II, and urinary AGT excretion in the ATS group were increased significantly at day 7 compared with the control group. Administration of olmesartan (ATS+ARB group) significantly decreased the levels of renal lesions, proteinuria, and intrarenal RAS activity compared with the ATS group. In addition, the levels of urinary AGT excretion correlated with the levels of glomerular damage, urinary protein excretion, and immunoreactivity for AGT and ANG II in kidney. On the other hand, plasma renin activity was significantly lower in the ATS group compared with the control group and significantly higher in the ATS+ARB group than in the ATS group. These data suggest that an increase in kidney-specific RAS activity, which parallels urinary AGT excretion, plays an important role in the development of ATS nephritis.

Increased urinary excretion of angiotensinogen(AGT) in type II diabetic mice

Diabetic nephropathy is the leading cause of end‐stage renal disease in the Western world leading to increased morbidity and mortality in type II diabetic patients. The renin‐angiotensin system (RAS) remains the major therapeutic target to slow the progression of renal disease in diabetes. The db/db mouse develops significant proteinuria, hyperglycemia, glomerulosclerosis, and insulin resistance with abnormalities similar to Type II diabetic nephropathy in humans. Ye et al. (Hypertension, 2004) reported that renal cortical angiotensin converting enzyme (ACE) activity is low, while ACE2 protein expression is high in db/db compared to db/m mice; therefore, studies were performed to test the hypothesis that intrarenal ANG II levels are reduced in db/db mice. Kidney ANG II (1,017±165 vs 788±99 fmol/g) and plasma ANG II levels (247±26 vs 391±105 fmol/ml) were not different between db/db and db/m mice, respectively. Kidney ANG I levels were also similar (113±24 db/db vs 110±45 fmol/g db/m; NS). In contrast, urinary excretion of AGT was significantly elevated in db/db compared to db/m mice (66±16 vs 12±2 ng/d, p&lt;0.001). Urinary excretion of albumin was also increased in db/db mice (1,075±170 vs 125±8; p&lt;0.001). Interestingly, excretion of AGT and albumin were positively correlated (R2=0.8). Polydipsia (7.8±1.1 vs 3.1±0.4 ml/d; p&lt;0.001), polyuria (5.1±0.9 vs 1.1±0.2; p&lt;0.001), hyperglycemia (408±19 vs 150±6 mg/dl; p&lt;0.001), hyperleptinemia (108.1±26 vs 4.2±0.8 mg/L; p&lt;0.001), and hyperinsulinemia (11.8±3.3 vs 1.3±0.4 mg/L; p&lt;0.001) are clearly evident in 18 wk old male db/db compared to db/m mice, respectively. In summary, urinary excretion of AGT and albumin are positively correlated and may represent a novel marker for the progression of diabetic renal disease.

Effects of Resibufogenin in Experimental Hypertension

Background/Aims: There are two major pathophysiologic processes involved in the development of hypertension: (1) expanded extracellular fluid volume and (2) vasoconstriction. We have developed a model of preeclampsia in the rat, in which excessive volume expansion (VE) plays a role. These animals excrete increased amounts of the bufodienolide, marinobufagenin (MBG), even before their hypertension and proteinuria become established. Furthermore, their hypertension is corrected by administration of resibufogenin (RBG), a compound structurally similar to MBG. Method: We studied two models of experimental hypertension in the nonpregnant animal, produced either by deoxycorticosterone acetate (DOCA)-salt administration or by angiotensin infusion. Results: RBG administered to the DOCA-salt rats lowered blood pressure and reduced proteinuria in the VE animals, but had no affect on the rats infused with angiotensin. Furthermore, although the production of superoxide anion in the aortas of both groups of hypertensive rats was increased over control, RBG reduced these levels to normal in the VE (DOCA-salt) animals only. RBG had no effect in the angiotensin-infused rats. The urinary excretion of angiotensinogen did not rise in VE-mediated hypertension, but did increase in the angiotensin-infused rats. Conclusions: MBG plays an important role in the causation of hypertension in the VE rats, but not in the vasoconstrictive model. RBG is effective only in VE-mediated hypertension.

Quantification of human angiotensinogen by a novel sandwich ELISA

The urinary angiotensinogen excretion rates show a clear relationship to kidney angiotensin II content, suggesting that urinary angiotensinogen may serve as an index of angiotensin II-dependent hypertensive rats. However, simple and accurate methods to measure human angiotensinogen are unavailable at this time. We have developed two antibodies and a sensitive and specific quantification ELISA system for human angiotensinogen to be applicable to human subjects. The ELISA is able to detect human angiotensinogen at range of 0.01–1 μg/well ( R 2 = 0.9945) using standard ELISA plates. This ELISA will be a useful tool to investigate the relationship between urinary angiotensinogen excretion rates and reactivity to antihypertensive drugs in hypertensive human subjects.

Expression of angiotensinogen in proximal tubule as a function of glomerular filtration rate

Proximal tubule (PT) angiotensinogen (AGT) is part of a tubular renin-angiotensin system (RAS) that participates in the regulation of sodium reabsorption along the entire nephron. Physiologic maneuvers affecting AGT expression in PT also affect systemic RAS. Here, we tested the hypothesis that PT AGT is regulated by increased glomerular filtration rate (GFR). Complete unilateral nephrectomy (UNX) in mice was used to induce a sustained increase in GFR in the remaining kidney. AGT expression was monitored by quantitative reverse transcription-polymerase chain reaction (RT-PCR). AGT protein in PT was investigated by semiquantitative histology. We also measured AGT concentration in plasma and in 24-hour urine by a specific enzyme-linked immunosorbent assay (ELISA). Seven weeks after nephrectomy, UNX animals exhibited a 2-fold increase in tubular AGT mRNA (P <.001) compared with sham-operated control animals. The proportion of PT sections exhibiting AGT immunostaining was significantly increased at day 3 (P <.05), and remained elevated at seven weeks (UNX = 0.63 +/- 0.09, sham = 0.38 +/- 0.02, P <.01), revealing recruitment of AGT-producing cells along the PT. AGT excretion in final urine corrected for creatinine and kidney weight was also elevated by UNX at seven weeks (UNX = 209 +/- 42 pmol/mg/g, sham = 147 +/- 29 pmol/mg/g, P <.05), with no difference in plasma AGT between UNX and control animals. These observations suggest that AGT expression in PT adapts in the long-term to changes in GFR. In the UNX model, urinary AGT excretion is also elevated as a consequence of increase in net tubular flow.

Intrarenal angiotensin II and hypertension.

Elevations in intrarenal angiotensin II (Ang II) cause reductions in renal function and sodium excretion that contribute to progressive hypertension and lead to renal and vascular injury. Augmentation of intrarenal Ang II occurs by several processes, leading to levels much greater than can be explained from the circulating levels. In Ang II-dependent hypertension, Ang II is internalized via an AT1 receptor mechanism, but there is also sustained intrarenal production of Ang II. Ang II exerts a positive feedback action on intrarenal angiotensinogen (AGT) mRNA and protein. The increased intrarenal AGT production is associated with increased intrarenal and intracellular Ang II contents and urinary AGT excretion rates. The increased urinary AGT indicates spillover of AGT into distal nephron segments supporting enhanced distal Ang II formation and sodium reabsorption. The augmentation of intrarenal Ang II provides the basis for sustained actions on renal function, sodium excretion, and maintenance of hypertension.

Enhancement of Intrarenal Angiotensinogen in Dahl Salt-Sensitive Rats on High Salt Diet

This study was performed to examine whether there is an inappropriate regulation of intrarenal angiotensinogen in Dahl-salt sensitive rats (DS) fed a high salt diet (HS). Dahl salt-resistant rats (DR) and DS were maintained on HS (8% NaCl) or low salt diet (LS, 0.3% NaCl) for 4 weeks. Systolic blood pressure (SBP), measured by tail-cuff plethysmography, was unaltered in DR (DR+HS, 127+/-3 mm Hg, n=5; DR+LS, 126+/-3, n=5); however, SBP was significantly increased in DS+HS (208+/-7, n=9) compared with DS+LS (134+/-2, n=5). HS suppressed plasma renin activity in both strains (0.7+/-0.2 ng of angiotensin I/mL per hour in DS+HS, 3.1+/-0.5 in DS+LS, 0.8+/-0.2 in DR+HS, 5.1+/-0.7 in DR+LS). Plasma angiotensinogen levels, measured by Western blot analysis, were also suppressed by HS in both strains (36 919+/-2170 integrated densitometric unit in DS+HS, 53 028+/-2752 in DS+LS, 44 722+/-1721 in DR+HS, 55782+/-3785 in DR+LS). However, kidney angiotensinogen levels were significantly increased in DS+HS (75 850+/-4171, integrated densitometric unit) compared with DS+LS (47 232+/-3470), DR+HS (44 748+/-8236), and DR+LS (42 504+/-4052). Urinary excretion of angiotensinogen, measured by radioimmunoassay of angiotensin I after incubation with excess renin, had a similar profile. Urinary excretion of angiotensinogen was significantly increased in DS+HS (2958+/-531 pmol/d) compared with DS+LS (56+/-4), DR+HS (31+/-12), and DR+LS (21+/-7). These data indicate that intrarenal angiotensinogen is enhanced in DS+HS, which is reflected by the increased urinary excretion of angiotensinogen. The results suggest that DS on HS have an inappropriate augmentation of intrarenal angiotensinogen, which may contribute to impaired sodium excretion during a high salt diet and the development of hypertension in this strain.