Proceedings of the Symposium ‘Angiotensin AT1 Receptors: From Molecular Physiology to Therapeutics’: STRUCTURAL CHANGES IN THE RENAL VASCULATURE IN THE SPONTANEOUSLY HYPERTENSIVE RAT: NO EFFECT OF ANGIOTENSIN II BLOCKADE

• 1 There is strong evidence for a renal basis to the development of hypertension in the spontaneously hypertensive rat (SHR). Alterations of the SHR renal vasculature, including the glomerulus, may be involved in the initiation and maintenance of hypertension in this animal model.
• 2 The arterial walls of pre-glomerular vessels of the SHR are hypertrophied compared with WKY vessels. Unlike other vascular beds in the SHR, this hypertrophy is independent of angiotensin II (AngII).
• 3 Glomerular number and volume are similar between SHR and the normotensive Wistar-Kyoto (WKY) rats. These results provide no support for the theory that a reduced filtration surface area within the kidneys of the SHR contributes to the elevated blood pressure in these animals.
• 4 Intrarenal hypertrophy may have similar haemodynamic consequences to clipping of the main renal artery, as in Goldblatt hypertension. Further analysis of the role of pre-glomerular arterial hypertrophy is warranted to determine its involvement in the initiation and maintenance of hypertension in the SHR.

     

Prevention of diabetes-induced albuminuria in transgenic rats overexpressing human aldose reductase

Studies using pharmacologic inhibitors have implicated the enzyme aldose reductase in the pathogenesis of albuminuria and diabetic renal disease. However, a clear conclusion is not easily drawn from such studies since these pharmacologic inhibitors have nonspecific properties. To examine further the role of aldose reductase, we have overexpressed the human enzyme in a transgenic rat model. Transgene expression in the kidney was predominantly localized to the outer stripe of the outer medulla, compatible with the histotopography of the straight (S3) proximal tubule. The effect of enzyme overexpression on diabetes-induced renal function and structure was then investigated. Contrary to what may have been anticipated from the previous enzyme inhibition studies, diabetes-induced albuminuria was completely prevented by the overexpression of aldose reductase. No effect of overexpression of aldose reductase on renal structure nor on urinary excretion of β₂-microglobulin and N-acetyl-β-d-glucosaminidase was observed in this transgenic rat model. In conclusion, our study strongly suggests that multiple roles for aldose reductase may give it a more complex place in diabetic nephropathy than is currently recognized.     

Renal and Cardiovascular Effects of Renal Medullary Damage with Bromoethylamine in Dogs

Bromoethylamine (BEA, 30-40 mg/kg) was administered to dogs to determine whether damage to the inner medulla of the kidney, the putative source of a depressor hormone, causes hypertension in this species. Bromoethylamine produces hypertension in rats but this has not been confirmed in other species, although we have shown that this dose of BEA in dogs abolishes the release of a reno-medullary vasodepressor hormone in response to marked increases in renal perfusion pressure. During acute BEA administration over 1 h to conscious dogs, there were no significant effects on renal blood flow, arterial pressure or total peripheral resistance, but there was a significantly greater diuresis compared to vehicle administration. Over the first 10-14 days after BEA, daily urine output rose 5-10 fold initially and plasma creatinine concentration rose markedly. There was no significant effect on arterial pressure, cardiac output, total peripheral resistance, or renal blood flow over this period. BEA administration caused extensive damage to the thin limbs of the loops of Henle, widespread thrombosis of blood vessels and haemorrhage into the interstitium of the dog renal medulla. Reno-medullary interstitial cells were devoid of lipid droplets, were synthetic, and were associated with increased amounts of extracellular matrix. Thus extensive renal medullary damage by BEA administration to conscious dogs did not alter resting systemic haemodynamics, and these results therefore provide no evidence for a role for the medulla in the maintenance of resting arterial pressure in the dog.     

Expression of transforming growth factor-β type II receptor mRNA in embryonic and adult rat kidney

Summary: The transforming growth factor-β (TGF-β) family of growth factors regulates cell proliferation, differentiation, extracellular matrix synthesis and angiogenesis in many developing tissues. Transforming growth factor-β1 was recently shown to affect the branching of ureteric epithelium and nephron formation in cultured rat metanephroi. As the TGF-β type II receptor is specific for the TGF-β family, the present study used in situ hybridization to localize mRNA for this receptor in metanephroi from Sprague-Dawley rat embryos. Transforming growth factor-β type II receptor mRNA was located in ureteric duct epithelium, undifferentiated mesenchymal cells in the nephrogenic zone, vesicles, comma-shaped bodies and S-shaped bodies. In some S-shaped bodies, TGF-β type II receptor mRNA was not expressed in the lower limb, which subsequently forms the renal corpuscle. Expression was not observed in capillary loop stage glomeruli and maturing glomeruli, or in proximal tubules and interstitial cells. In adult rat kidney, TGF-β type II receptor mRNA was expressed in cortical collecting ducts and distal tubules but not in glomeruli or proximal tubules. These findings demonstrate that the prominent expression of TGF-β type II receptor mRNA decreases as glomeruli and tubules develop. Expression then remains undetectable in adult glomeruli and proximal tubules. the developmentally-regulated expression of this receptor suggests a key role in glomerular and nephron development.     

Glomerular stereology: Why, what and how to measure glomerular structure

Summary: Quantitative methods are frequently used to analyse the structure of renal glomeruli. However, on most occasions, measurements are made on glomerular profiles (the two-dimensional samples of glomeruli seen in histological sections), and provide little or no information about the structure of whole, three-dimensional glomeruli. Stereology is the discipline concerned with the quantitative analysis of three-dimensional structures. With stereology one can estimate the total number of glomeruli in kidneys, as well as mean glomerular volume, the number of cells in glomeruli, and the length and surface area of glomerular capillaries. In addition to providing a means for detecting structural differences between glomeruli from different specimens, stereology provides quantitative structural information that can be correlated with quantitative physiological, biochemical and molecular data. Over the past decade we have witnessed the development of a new generation of unbiased, cost-efficient stereological methods that are ideally suited to analysing glomeruli. Some of these methods are introduced in this review, and then three recent studies from our laboratories that successfully utilized these methods are described. These studies concerned hypertension, kidney development, and the pathogenesis of focal and segmental glomerulosclerosis.     

Immunolocalization of fibroblast growth factor-1 and -2 in the embryonic rat kidney

Summary: Fibroblast growth factors (FGF) regulate cell proliferation, migration, differentiation and angiogenesis during morphogenesis in many different tissues. Recent evidence indicates that exogenous FGF-2 stimulates mesenchymal condensation in cultured rat metanephroi, a crucial epithelial-mesenchymal induction event in the developing nephron. the aim of the present investigation was to determine the in vivo distribution of FGF-1 and FGF-2 in developing rat metanephroi at embryonic days 14, 15, 16, 18 and 20. Avidin-biotin enhanced indirect immunohistochemistry was used to demonstrate that both FGF-1 and FGF-2 were co-localized in metanephroi at all ages studied. High levels of FGF-1 and FGF-2 were present in ureteric bud branches and in developing distal tubules. Fibroblast growth factor-1 and FGF-2 were colocalized in developing nephron elements, from vesicles to S-shaped bodies, and in the mesangium of capillary loop and maturing stage glomeruli. Both growth factors were present in the mesenchyme of the nephrogenic zone and in the interstitium of the developing cortex. However, immunostaining for FGF was not evident in mesenchymal condensates, endothelial cells, medullary interstitial cells, or in the thin undifferentiated epithelium of the immature loop of Henle. These findings indicate that the expression of both FGF-1 and FGF-2 is tightly regulated in the embryonic kidney and suggest a role for these molecules in kidney development.     

Proceedings of the Symposium ‘Angiotensin AT1 Receptors: From Molecular Physiology to Therapeutics’: PRESENCE OF ANGIOTENSIN II AT2 RECEPTOR BINDING SITES IN THE ADVENTITIA OF HUMAN KIDNEY VASCULATURE

• 1 Angiotensin II (AngII) receptor subtypes in adult human kidney were pharmacologically characterized by in vitro autoradiography using the AngII receptor subtype-selective antagonists, losartan and PD 123319, and the sensitivity to the reducing agent, dithiothreitol.
• 2 High densities of AngII AT₁ receptor binding occur in the glomeruli and the inner stripe of the outer medulla, while a moderate AT₁ receptor binding is localized in the proximal convoluted tubules.
• 3 AT₂ receptor binding is observed predominantly in the intrarenal large blood vessels, including the arcuate, inter- and intra-lobular arteries, and in the renal capsule.
• 4 In the major renal artery, AT₁ receptor binding is abundant in the media and adventitia, while AT₂ receptor binding is observed mainly in the adventitia.
• 5 At the light microscopic level using emulsion autoradiography, AT₁ receptors are localized in the glomeruli and juxtaglomerular apparatus, as expected. However, in larger renal blood vessels, including the arcuate arteries, inter- and intra-lobular arteries, intense AT₂ receptor labelling occurs primarily in the adventitia, while the endothelium and vascular smooth muscle layers contain only low levels of AngII receptor binding.
• 6 These results indicate that the adult human kidney displays two pharmacologically distinct AngII receptor subtypes, with AT¹ predominating in the glomeruli, juxtaglomerular apparatus, proximal tubules and the inner stripe of the outer medulla, while AT₂ predominates in the adventitia of the arcuate and interlobular arteries and the renal capsule. The functional significance of AT₂ receptor binding sites in the adventitia of adult human kidney vessels remains to be elucidated.