Colocalization and release of angiotensin and renin in renal cortical cells

Abstract

Angiotensin is generated within the kidney, but the precise loci for the formation of angiotensin I (ANG I) and angiotensin II (ANG II) have not been demonstrated. We performed electron microscopy immunocytochemistry in kidney sections of 10-day-old (newborn) and adult Wistar-Kyoto (WKY) rats using specific antibodies to renin, ANG I, ANG II, and angiotensinogen (AO). Renin, ANG I, ANG II, and AO were present in juxtaglomerular (JG) cells. Renin was largely confined to cytoplasmic granules; ANG I and ANG II were colocalized to these granules but also were present in the cytoplasm; AO was distributed throughout the cytoplasm. AO also was present in a renal cortical distribution in proximal tubular cells. Northern blot analysis demonstrated AO mRNA in total kidney and liver but not in renal microvessels. Using the reverse hemolytic plaque assay, we demonstrated release of ANG I and renin from individual renocortical cells of adult WKY rats. Under control conditions, the number of releasing cells was 11 +/- 1 for ANG I and 10 +/- 1 for renin. Addition of rat renin inhibitor (RI) (1 x 10(-5) M), which inhibited renin activity in the medium from 37 to 9 pg ANG I.ml-1.h-1, did not alter ANG I plaque number. Addition of rat AO increased ANG I plaque number to 17 +/- 2 (P less than 0.05). Incubation with both RI and AO prevented the increase in ANG I plaque number obtained with AO alone. Enalapril treatment (7 days; n = 5) increased the number of plaque-forming cells to 22 +/- 2 for ANG I (P less than 0.0005) and to 39 +/- 7 for renin (P less than 0.001). The results suggest an intracellular location for AO and angiotensin and release of renin and ANG I by renal cortical cells and suggest that released angiotensin is produced intracellularly and that secretion of ANG I is augmented by converting enzyme inhibition.

Biomechanical coupling in renin-releasing cells.

“…Cells were cultured for 48 h on Permanox plastic-coated multi-well chamber slides (Nunc, Inc., Naperville, IL). Immunoperoxidase staining was performed using the avidin-biotin peroxidase complex (ABC) kit from Vector Laboratories, Inc. (Burlingame, CA), as described previously (10,11). Cells were fixed in methanol (4 Њ C) for 10 min, followed by a 10-s exposure to acetone (4 Њ C), and then air dried.…”

Angiotensinogen localization and secretion in the rat pancreas

“…In particular, local RAS components or tissue generation of Ang II have been detected in several organs such as brain (Phillips et al 1993, Sernia 1995, pituitary (Thomas & Sernia 1990a), adrenal glands (Vinson 1995, Mulrow & Franco-Saenz 1996, heart (Dostal 2000), kidneys (Celio 1981, Hunt et al 1992, Darby & Sernia 1995, gonads (Thomas & Sernia 1990b, Vinson et al 1997) and pancreas (Leung et al 1998, 1999, Tahmasebi et al 1999, Leung & Carlsson 2001). The role played by these tissue RAS is not well understood and they have been proposed to regulate local blood flow or to fulfil specific functions according to the tissue.…”

Review: Intrarenal angiotensin II levels in normal and hypertensive states

“…32,33,53 In contrast to vascular smooth muscle cells, proximal [57][58][59][60] Immunohistochemical studies using antibodies to angiotensinogen have clearly localised angiotensinogen to the proximal tubule segment. 56,61 These results indicate that much of the intrarenally-produced angiotensinogen is derived from proximal tubule cells and suggest it is one source of the intratubular concentrations of Ang I and Ang II. It has been noted that the preponderance of immunoreactive angiotensinogen in proximal tubule cells is located along the luminal membrane of the proximal tubule cells thus supporting the notion that angiotensinogen could be secreted into the tubular lumen and/or producing its metabolites intracellularly and secreting them directly into the tubule lumen.…”

“…Studies utilising immunohistochemical techniques demonstrated that Ang I and Ang II are co-localised with renin in JGA cells and vascular smooth muscle cells of the afferent arteriole. [53][54][55][56] These studies suggested that Ang II could be formed or internalised into the smooth muscle cells. 32 Because there is no direct evidence for intracellular angiotensinogen in vascular smooth muscle cells, it is more likely that Ang I and/or Ang II are internalised via receptor-mediated endocytosis in smooth muscle cells.…”

“…It has been noted that the preponderance of immunoreactive angiotensinogen in proximal tubule cells is located along the luminal membrane of the proximal tubule cells thus supporting the notion that angiotensinogen could be secreted into the tubular lumen and/or producing its metabolites intracellularly and secreting them directly into the tubule lumen. 56,61,62 To test for the presence of angiotensinogen in proximal tubule fluid, proximal tubule fluid samples were collected and incubated with excess renin, in order to determine total available Ang II from angiotensinogen or related precursors secreted into the lumen. 63 As shown in Figure 3, we found that the proximal tubule fluid angiotensinogen concentrations in rats were over 300 nM and greatly exceeded the free Ang I and Ang II tubular fluid concentrations.…”