Intrarenal Renin-Angiotensin System: Locally Synthesized or Taken up via Endocytosis?

The renin-angiotensin system (RAS) has long been recognized to play a preeminent role in the control of blood pressure and the development of hypertension. There is a general consensus that both the circulating (endocrine) and local (paracrine) RAS act together to regulate vascular tone, sympathetic outflow, renal tubular and vascular function, pressure natriuresis and blood pressure. However, it has been very difficult to dissect the specific contributions of the circulating versus the local RAS to the regulation of renal function because components of the RAS in the proximal tubule can be absorbed and/or taken up by endocytosis as well as synthetized locally. In this issue of Hypertension, Roksner et al. used a variety of approaches to better define the origin of renin and prorenin found in the proximal tubule and urine.¹

The existence of the intrarenal RAS is strongly supported by the observations that the concentrations of angiotensin (Ang) I and II in proximal tubular fluid and in the kidney often exceed that of plasma, and components of the RAS in the kidney can be regulated independently from circulating levels.² In addition, prorenin, angiotensinogen (AGT) and angiotensin converting enzyme (ACE) are all synthetized in various nephron segments.² Indeed, as summarized in Figure 1, Ang I and II as well as renin, prorenin, AGT and ACE proteins are all expressed in the proximal tubule. Furthermore, prorenin and ACE proteins are localized in the cortical collecting duct. Prorenin, renin, AGT, and Ang II are also excreted and the levels in urine increase in some forms of hypertension and renal disease.^3,4 The prevailing view is that circulating Ang I and II are filtered and reabsorbed in the proximal tubule. Ang II is also formed locally through the interactions of renin, AGT and ACE in this portion of the nephron. Together, the filtered and locally produced Ang II regulates Na⁺ reabsorption in the proximal tubule (5–7). AGT with a molecular weight (MW) of 63 kD was originally thought to be too large to be filtered. This led to the proposal that most of the AGT in proximal tubular fluid was synthesized locally and secreted. It was further suggested that the secreted AGT later interacts with renin derived from prorenin secreted by principal cells in the collecting duct to form Ang II in the presence of ACE that is avidly expressed in this segment.² The Ang II that is formed is thought to enhance sodium transport in the collecting duct. Indeed, AGT, renin and Ang II are found in the urine of man and experimental animals. Moreover, these excreted proteins have been proposed to serve as biomarkers of the activity of the intrarenal RAS since urinary levels are elevated in many forms of hypertension and various renal disease models.^2,8

Figure 1.

Summary of the origin of components of the intrarenal renin-angiotensin system (RAS) found along the nephron. Ang I and Ang II (MW 1 kD), renin (MW 40 kD), prorenin (MW 44 kD), and angiotensinogen (AGT) (MW 63 kD) are filtered in the glomerulus. Ang I and II, renin, prorenin, and AGT are extensively reabsorbed in the proximal convoluted tubule. Angiotensin converting enzyme (ACE) is synthesized locally and abundantly expressed in apical membrane of the proximal tubule. AGT is also synthesized in the proximal tubule. Prorenin is synthesized and is found in storage granules in principal cells of the collecting duct. ACE is also expressed in this portion of the nephron. Prorenin is secreted by the principal cells especially in Ang II hypertensive animals, but it remains to be determined whether prorenin is converted to renin in the collecting duct. Prorenin, renin, AGT and Ang II are excreted in the urine and their levels are increased following the development of hypertension and in various renal diseases.

In this issue of Hypertension, Roksner et al.¹studied the filtration of fluorescently labeled renin and prorenin by the glomerulus using multiphoton microscopy in C57BL/6J mice before and after damaging the glomerular filtration barrier by treating the animals with doxorubicin. They found that the filtration of both renin (MW 40 kD) and prorenin (MW 44 kD) exceeded that of albumin (MW 67 kD) and both increased after administration of doxorubin. About 97% of the filtered renin was reabsorbed along the nephron, whereas the reabsorption of prorenin was complete. None was detected in the urine even after proteinuria was induced with doxorubin. In other studies, they measured the excretion of renin and prorenin in CYP1a1-Ren2 transgenic rats. Upregulation of the production of prorenin in the liver of these animals increased circulating levels of prorenin and renin by 200 and 20-fold, respectively. This resulted in a 1000-fold increase in urinary renin excretion, but surprisingly the reabsorption of prorenin remained complete, and none was detected in the urine. Finally, Roksner et al. measured the excretion of renin and prorenin in the urine of patients with Dent’s disease or Lowes syndrome which are associated with defective megalin-mediated reuptake of filtered proteins in the proximal tubule.¹ The urinary levels of renin, AGT and albumin in these patients were 20 to 40-fold higher than that reported in the urine of normal patients, and prorenin could be detected in the urine of these patients. These findings suggest that renin and prorenin are filtered by the glomerulus and extensively reabsorbed in the proximal tubule along with albumin via a megalin-dependent pathway. Prorenin could only be detected in the urine after megalin-mediated reuptake was impaired.

These results are consistent with a recent report that AGT (MW 63 kD) is filtered by about the same extent as albumin (67 kD) and is taken up in the early proximal tubule via the same megalin-dependent process.⁹ These authors further reported that the synthesis of AGT is restricted to the proximal straight tubule, suggesting that locally formed AGT probably does not contribute to the production of Ang I in the proximal tubule (9). A subsequent study by Masusaka et al.¹⁰ demonstrated that liver-specific knockout of AGT marked reduced intrarenal levels of AGT and Ang II, indicating that most of AGT in the kidney and intrarenally formed Ang II are dependent on the filtration and subsequent reuptake of circulating AGT rather than local synthesis.

The view that emerges from the current study of Roksner et al.¹ and previous findings of Pohl et al.⁹and Masusaka et al.¹⁰ is that most of the prorenin, renin and AGT in the urine (Figure 1) is filtered and escapes reabsorption in the proximal tubule. Likewise, urinary Ang II is derived from that which is filtered and escapes reabsorption and metabolism in the proximal tubule, as well as from Ang I formed from the interaction of filtered and/or reabsorbed AGT with renin in the lumen of the proximal tubule, which is converted to Ang II by ACE expressed along the brush border. The competing hypothesis is that a significant fraction of urinary Ang II is produced by renin secreted in the collecting duct that interacts with AGT in distal tubular fluid to form Ang I which is converted by ACE along the collecting duct.^2,8 Arguing against this possibility is the finding that the immunoreactive renin in storage granules in the collecting duct is in the form of prorenin. Thus, most of the renin found in the urine should be in the form of prorenin unless an efficient mechanism exists to activate prorenin to renin. However, in the current study, Roksner et al.¹ reported that they were unable to detect prorenin in the urine of rats or mice under a variety of experimental conditions.

The strongest argument for the independent regulation of the intrarenal RAS is that the levels of prorenin, renin, AGT, and Ang II are elevated in the kidney and urine of Ang II-infused animals when plasma renin activity is suppressed.^2,8 This led to the suggestion that the elevated circulating levels of Ang II augment the synthesis and secretion of AGT in the proximal tubule and prorenin/renin in the collecting duct to serve as a feed-forward mechanism to further increase the intrarenal formation of Ang II.^2,3 A similar activation of the intrarenal RAS system was also proposed in salt-sensitive hypertensive and diabetic animals in which circulating levels of renin are also suppressed.² In these models, increased levels of inflammatory cytokines and reactive oxygen species were suggested as the driving factors for activation of the intrarenal RAS.² The current findings of Roksner et al.¹ could also explain the apparent activation of the intrarenal RAS in hypertension, if one considers that the glomerular filtration barrier is likely damaged so that elevations in the filtration of renin, prorenin and AGT might stimulate the local formation of Ang II. The urinary excretion of prorenin, renin and AGT may also increase in hypertensive and renal disease models secondary to an increase in the filtered load of albumin which competes with megalin-dependent uptake of prorenin, renin and AGT in the proximal tubule.

In summary, the concept of a local intrarenal RAS remains firmly established despite the lack of consensus as to whether the components of this system are produced locally or filtered and reabsorbed along the nephron. There is no question that Ang II is generated in the kidney and plays an essential role in regulating sodium and water reabsorption, the pressure natriuretic relationship and the long term control of blood pressure. However, because prorenin, renin, AGT and Ang II are filtered and reabsorbed, it has been difficult to determine what fraction of these proteins in the proximal tubule is synthetized locally or taken up from tubular fluid. A previous study using a liver specific AGT KO mouse suggests that most of the AGT in the kidney is filtered and reabsorbed¹⁰ and the current study suggests that the same is true for renin and prorenin found in the proximal tubule.¹ It remains to be determined if renin and AGT taken up and stored in endosomes can be secreted back into tubular fluid. Additional studies are also needed to determine whether prorenin and renin in distal tubular fluid and urine are filtered and escapes reabsorption in the proximal tubule or are synthesized and secreted in the collecting duct. The percentage of urinary Ang II formed in the proximal tubule versus the collecting duct also remains to be determined, especially in man. Clinically, the results of the present¹ and past studies^9,10suggest that increases in renin and AGT excretion in hypertensive and diabetic patients (8) might reflect changes in filtration and tubular reabsorption rather than serve as biomarkers for activation of the intrarenal RAS. Finally, the clinical significance of the studies on the origin of components of the intrarenal RAS remains uncertain, since most of the data is based on animal studies. Moreover, the beneficial effects of ACE and renin inhibitors to block the intrarenal formation of Ang II, or ARBs to inhibit renal AT1 receptors, probably do not depend on whether the components of this system are synthesized locally or filtered and reabsorbed.